Non-nucleotide composition and method for inhibiting platelet aggregation

ABSTRACT

This invention is directed to a method of preventing or treating diseases or conditions associated with platelet aggregation. The method is also directed to a method of treating thrombosis or related disorders. The method comprises administering to a compounds useful for this invention include compounds of general Formulae I and III-XII, or salts, hydrates, and solvates thereof. The present invention also provides novel compounds of Formulae I and III-XII.

This application is a continuation-in-part of U.S. application Ser. No.10/971,766, filed Oct. 21, 2004, which claims priority to U.S.provisional application No. 60/513,845 filed Oct. 21, 2003. The contentsof both applications are incorporated herein by reference in theirentirety.

TECHNICAL FIELD

This invention relates to non-nucleotide compounds and methods of makingand using such compounds in the prevention or treatment of diseases orconditions associated with platelet aggregation, including thrombosis,stroke and myocardial infarction in humans and other mammals, and forinhibition of platelet aggregation in blood and blood-related products.

BACKGROUND OF THE INVENTION

Hemostasis is the spontaneous process of arresting bleeding from damagedblood vessels. Upon injury, precapillary vessels contract withinseconds, and thrombocytes, or blood platelets, bind to the exposedsubendothelial matrix of an injured vessel by a process called plateletadhesion. Platelets also stick to each other in a phenomenon known asplatelet aggregation to form stable platelet aggregates that quicklyhelp stop or slow blood outflow from injured vessels.

An intravascular thrombus can result from pathological disturbances ofhemostasis, or by the rupture of atherosclerotic plaques. Plateletadhesion and aggregation are critical events in intravascularthrombosis. Activated under conditions of high shear blood flow indiseased vessels or by the release of mediators from other circulatingcells and damaged endothelial cells lining the vessel, platelets andother cells accumulate at a site of vessel injury to form a thrombus,and recruit more platelets to the developing thrombus. The thrombus cangrow to sufficient size to block off arterial blood vessels. Thrombi canalso form in areas of stasis or slow blood flow in veins. Venous thrombican easily detach portions of themselves, creating emboli that travelthrough the circulatory system. This process can result in blockade ofother vessels, such as pulmonary arteries. Blockages of this sort canresult in pathological outcomes such as pulmonary embolism. Thus,arterial thrombi cause serious disease by local blockade, whereas themorbidity and mortality associated with venous thrombi arise primarilyafter distant blockade, or embolization. Conditions associated withpathological thrombus formation include venous thromboembolism,thrombophlebitis, deep vein thrombosis, arterial embolism, coronary andcerebral arterial thrombosis, unstable angina, myocardial infarction,stroke, transient ischemic attack, cerebral embolism, renal embolism andpulmonary embolism.

A number of converging pathways lead to platelet aggregation. Whateverthe initial stimulus, the final common event is crosslinking ofplatelets by binding of fibrinogen to a membrane binding site,glycoprotein IIb/IIIa (GP IIb/IIIa, also known as integrin α_(IIb)β₃).Antagonists of the GP IIb/IIIa receptor have been shown to producepotent antithrombotic effects (Ali, U.S. Pat. No. 6,037,343; Duggan, etal., U.S. Pat. No. 6,040,317). GP IIb/IIIa antagonists includefunction-blocking antibodies like Abciximab (ReoPro®), cyclic peptidesand peptidomimetic compounds (The EPIC investigators; Califf, R. M.,coordinating author, New Engl. J. Med. 330: 956-961 (1994); TheIMPACT-II investigators, Lancet 349:1422-1428 (1997); The RESTOREinvestigators, Circulation 96: 1445-1453 (1997)). The clinical efficacyof some of these newer drugs, such as Abciximab, is impressive, butrecent trials have found that these approaches are associated with anincreased risk of major bleeding, sometimes necessitating bloodtransfusion (The EPIC investigators; Califf, R. M., coordinating author,New Engl. J. Med. 330: 956-961 (1994)). Also, administration of thisclass of antiplatelet agent appears to be limited to intravenousmethods.

Thrombin can produce platelet aggregation independently of otherpathways but substantial quantities of thrombin are unlikely to bepresent without prior activation of platelets by other mechanisms.Thrombin inhibitors, such as hirudin, are highly effectiveantithrombotic agents. However, functioning as both antiplatelet andanti-coagulant agents, thrombin inhibitors again can produce excessivebleeding (The TIMI 9a Investigators, Circulation, 90: 1624-1630 (1994);The GUSTO IIa Investigators, Circulation, 90: 1631-1637 (1994); Neuhaus,et al., Circulation, 90: 1638-1642 (1994)).

Various antiplatelet agents have been studied as inhibitors of thrombusformation. Some agents such as aspirin and dipyridamole have come intouse as prophylactic antithrombotic agents, and others have been thesubjects of clinical investigations. To date, therapeutic agents such asthe disintegrins, and the thienopyridines ticlopidine (TICLID®) andclopidogrel (PLAVIX®) have been shown to have utility as plateletaggregation inhibitors, although they can produce a substantial numberof side effects and have limited effectiveness in some patients. (Hass,et al., N. Engl. J. Med., 321: 501-507 (1989); Weber, et al., Am. J.Cardiol. 66: 1461-1468 (1990); Lekstrom and Bell, Medicine 70: 161-177(1991)). In particular, the use of the thienopyridines in antiplatelettherapies has been shown to increase the incidence of potentially lifethreatening thrombotic thrombocytopenic purpura (Bennett, et al., N.Engl. J. Med, 342: 1771-1777 (2000)). Aspirin, which has a beneficialeffect on the inhibition of platelet aggregation (AntiplateletTrialists' Collaboration, Br. Med. J. 308: 81-106 (1994); AntiplateletTrialists' Collaboration, Br. Med. J. 308: 159-168 (1994)), acts byinhibiting the synthesis of prostaglandins. Its well-documented, highincidence of gastric side effects, however, limits its usefulness inmany patients. In addition, aspirin resistance has been observed in someindividuals (McKee, et al., Thromb. Haemost. 88: 711-715 (2002)).

Many studies have demonstrated that adenosine 5′-diphosphate (ADP) playsa key role in the initiation and progression of arterial thrombusformation (Bernat, et al., Thromb. Haemostas. 70: 812-826 (1993));Maffrand, et al., Thromb. Haemostas. 59: 225-230 (1988); Herbert, etal., Arterioscl. Thromb. 13: 1171-1179 (1993)). ADP induces inhibitionof adenylyl cyclase and modulation of intracellular signaling pathwayssuch as activation of phosphoinositide-3 kinase (PI3K), influx andmobilization of intracellular Ca⁺², secretion, shape change, andplatelet aggregation (Dangelmaier, et al. Thromb Haemost. 85: 341-348(2001)). ADP-induced platelet aggregation is triggered by its binding tospecific receptors expressed in the plasma membrane of the platelet.There are at least three different P2 receptors expressed in humanplatelets: P2X₁, P2Y₁, and P2Y₁₂. The P2X₁ receptor is a ligand-gatedcation channel that is activated by ATP, resulting in a transient influxof extracellular calcium. This receptor has been implicated in theregulation of platelet shape change, and recent evidence suggests itsparticipation in thrombus formation in small arteries under high shearforces. (Jagroop, et al., Platelets 14:15-20 (2003); Hechler, et al., J.Exp. Med. 198: 661-667 (2003)). The P2Y₁ receptor is a G protein-coupledreceptor that is activated by ADP, and is responsible for calciummobilization from intracellular stores, platelet shape change andinitiation of aggregation. The P2Y₁₂ receptor, also referred to as theP2Y_(ac) and P2_(T) receptor, is a G protein-coupled receptor that isactivated by ADP and is responsible for inhibition of adenylyl cyclaseand activation of PI3K. Activation of P2Y₁₂ is required for plateletsecretion and stabilization of platelet aggregates (Gachet, Thromb.Haemost. 86: 222-232 (2001); André, et al., J. Clin. Invest., 112:398-406 (2003)).

ADP-induced platelet aggregation requires the simultaneous activation ofboth P2Y₁ and P2Y₁₂ receptors, and therefore, aggregation can beinhibited by blockade of either receptor. Several authors havedemonstrated that ADP-induced aggregation is inhibited in aconcentration-dependent manner by analogues of adenosine triphosphate(ATP). ATP, itself, is a weak and nonselective, but competitive, P2Y₁and P2Y₁₂ receptor antagonist. Ingall, et al. (J. Med. Chem. 42: 213-220(1999)) have reported that modification of the polyphosphate side chainof ATP along with substitution of the adenine moiety at the C²-position,resulted in compounds that inhibited the P2_(T) receptor (or P2Y₁₂receptor). Zamecnik (U.S. Pat. No. 5,049,550) has disclosed a method forinhibiting platelet aggregation by administration of a diadenosinetetraphosphate-like compound, App(CH₂)ppA. Kim and Zamecnik (U.S. Pat.No. 5,681,823) have disclosed P¹, P⁴-(dithio)-P²,P³-(monochloromethylene)-5′, 5′″-diadenosine-P¹, P⁴-tetraphosphate as anantithrombotic agent.

Nucleotide P2Y₁₂ antagonists have been developed, however, there isstill a need for compounds that have improved oral bioavailability andblood stability.

Thienopyridines, ticlopidine and clopidogrel react covalently with theP2Y₁₂ receptor and produce irreversible platelet inhibition in vivo(Quinn and Fitzgerald, Circulation 100: 1667-1672 (1999); Geiger, etal., Arterioscler. Thromb. Vasc. Biol. 19: 2007-2011 (1999); Savi, etal., Thromb Haemost. 84: 891-896 (2000)). Patients treated withthienopyridines usually require 2-3 days of therapy to observesignificant inhibition of platelet aggregation, however, and maximalinhibition usually is observed between 4 to 7 days after initiation oftreatment. Also, the platelet inhibitory effect of thienopyridinespersists up to 7-10 days after the therapy is discontinued, and bothticlopidine and clopidogrel produce a significant prolongation of thebleeding time (from 1.5 to 2-fold over control). Because of theprolonged effect of thienopyridines, these drugs need to be discontinuedfor 7 to 10 days prior to elective surgery, leaving the patientunprotected from a possible thrombotic event during that period.Recently, the association of thienopyridine treatment with events ofthrombotic thrombocytopenic purpura has been reported (Bennett, et al.,N. Engl. J. Med. 342: 1773-1777 (2000); Bennett, et al., Ann. Intern.Med. 128: 541-544 (1998)).

Derivatives of5,7-disubstituted-1,2,3-triazolol[4,5-d]pyrimidin-3-yl-cyclopentanes and-tetrahydrofurans have been disclosed as antagonists of the P2T- (orP2Y₁₂) receptor on platelets (Cox, et al., U.S. Pat. No. 5,747,496, andrelated patents; Bonnert, et al., U.S. Pat. No. 6,297,232; WO 98/28300;Brown, et al., WO 99/41254; WO 99/05144; Hardern, et al. WO 99/05142; WO01/36438; and Guile, et al. WO 99/05143) for use in the treatment ofplatelet aggregation disorders.

Guile, et al. (WO 00/04021) disclose the use oftriazolo[4,5-d]pyrimidine compounds in therapy. Brown, et al. (U.S. Pat.No. 6,369,064) disclose the use of Triazolo(4,5-d)pyrimidine compoundsin the treatment of myocardial infarction and unstable angina. Dixon, etal. (WO 02/096428) disclose the use of 8-azapurine derivatives incombination with other antithrombotic agents for antithrombotic therapy.Springthorpe discloses AZD6140 as a potent, selective, orally activeP2Y₁₂ receptor antagonist which is now in Phase I clinical trials(Abstracts of Papers, 225^(th) ACS National Meeting, New Orleans, La.;March, 2003; MEDI-016). WO 02/016381 discloses a method of preventing ortreating diseases or conditions associated with platelet aggregationusing mononucleoside polyphosphates and dinucleoside polyphosphates.

There is still a need in the areas of cardiovascular and cerebrovasculartherapeutics, and in blood product preparation, purification, andstorage, for selective, reversible inhibitors of platelet activation,which can be used in the prevention and treatment of thrombi or otheraggregation-related problems.

SUMMARY OF THE INVENTION

This invention is directed to methods of preventing or treating diseasesor conditions associated with platelet aggregation or where theaggregation of platelets inhibits treatment options. This invention isdirected to methods of preventing or treating thrombosis and relateddisorders. This invention is further directed to methods of inhibitingplatelet aggregation in blood and blood products comprising platelets,such as stored blood.

The method comprises administering to a mammalian subject or to a samplecomprising blood or platelet-comprising material, a compositioncomprising one or more non-nucleotide P2Y₁₂ receptor antagonist compoundthat effectively binds to P2Y₁₂ receptors on platelets, preferably in areversible manner, and thereby causes an inhibition of the ADP-inducedplatelet aggregation response in blood or in a platelet-comprisingmaterial. The compounds useful for the methods are compounds of generalFormula I, III-XII, and/or tautomers thereof, and/orpharmaceutically-acceptable hydrates, solvates, and/or salts thereof.

The invention also provides novel compounds and pharmaceuticalcompositions. The compounds of Formulae I, and III-XII are useful inthat they possess antagonist activity at platelet P2Y₁₂ receptors.

Optionally, the compounds of this invention can be used in combinationwith other compounds useful for the treatment of platelet aggregationdisorders or diseases.

DETAILED DESCRIPTION OF THE INVENTION

Definitions

When present, unless otherwise specified, the following terms aregenerally defined as, but are not limited to, the following:

Alkyl groups are from 1 to 12 carbons inclusively, either straightchained or branched, with or without unsaturation and with or withoutheteroatoms, are more preferably from 2 to 8 carbons inclusively, andmost preferably 2 to 6 carbons inclusively.

Alkenyl groups are from 1 to 12 carbons inclusively, either straight orbranched containing at least one double bond but may contain more thanone double bond, with or without heteroatoms.

Alkynyl groups are from 1 to 12 carbons inclusively, either straight orbranched containing at least one triple bond but may contain more thanone triple bond, and additionally may contain one or more double bondedmoieties, with or without heteroatoms.

Cycloalkyl groups from 3 to 12 carbons inclusively, more preferably from3 to 10 carbons inclusively, and most preferably 3 to 6 carbonsinclusively, with or without unsaturation, and with or withoutheteroatoms.

Aralkyl groups are from 1 to 8 carbons inclusively in the alkyl portion,are more preferably from 1 to 6 carbons inclusively in the alkylportion, and most preferably are 1 to 4 carbons inclusively in the alkylportion; as included in the alkyl definition above, the alkyl portion ofan aralkyl group can include one or more positions of unsaturation suchas a double bond or a triple bond in the chain when the chain includestwo or more carbon atoms; the alkyl portion of an aralkyl group can alsoinclude one or more heteroatoms and/or substituents; the aryl portion ofan aralkyl group can be a monocyclic or polycyclic moiety from 3 to 8carbons inclusively per ring in the aryl portion, more preferably from 4to 6 carbons inclusively per ring, and most preferably 5 to 6 carbonsinclusively per ring; the aryl portion of an aralkyl group can also bearone or more substituents and/or heteroatoms.

Aryl groups are either monocyclic or polycyclic, are from 3 to 8 carbonsinclusively per ring, are more preferably from 4 to 6 carbonsinclusively per ring, and are most preferably 5 to 6 carbons inclusivelyper ring; aryl groups can also bear substituents and/or heteroatoms.

Heteroaralkyl groups are from 1 to 8 carbons inclusively in the alkylportion, are more preferably from 1 to 6 carbons inclusively in thealkyl portion, and most preferably are 1 to 4 carbons inclusively in thealkyl portion; as included in the alkyl definition above, the alkylportion of a heteroaralkyl group can include one or more positions ofunsaturation such as a double bond or a triple bond in the chain whenthe chain includes two or more carbon atoms; the alkyl portion of aheteroaralkyl group can also include one or more heteroatoms and/orsubstituents; the heteroaryl portion of a heteroaralkyl group can be amonocyclic or polycyclic moiety from 3 to 8 carbons inclusively per ringin the heteroaryl portion and containing from 1 to 4 heteroatomsinclusively per ring, more preferably from 4 to 6 carbons inclusivelyper ring, and most preferably 5 to 6 carbons inclusively per ring; theheteroaryl portion of an heteroaralkyl group can also bear one or moresubstituents and/or heteroatoms.

Heteroaryl groups are either monocyclic or polycyclic, contain from 1 to4 heteroatoms inclusively per ring, are from 3 to 8 atoms inclusivelyper ring, are more preferably from 4 to 6 atoms inclusively per ring,and are most preferably 5 to 6 atoms inclusively per ring; heteroarylgroups can also bear substituents and/or heteroatoms.

Substituents on the foregoing groups can be, but are not limited to,hydroxy, nitro, methoxy, fluoro, chloro, bromo, iodo, methyl, ethyl,propyl, butyl, thioalkyl, alkoxy, carboxyl, carboxamido, alkylsulfonyl,alkylsulfonylamino, sulfonamido, cyano, amino, substituted amino,trifluoromethyl, trifluoromethoxy, phenyl, pyridyl, imidazolyl,cyclopropyl, cyclopentyl, and cyclohexyl; and preferred heteroatoms areoxygen, nitrogen, and sulfur.

A desired substituent on a chain or ring (in place of a hydrogen at aposition) is one selected from the given alkyl, aryl, halogen, aralkyl,carboxy, alkoxycarbonyl, hydroxyl, acyloxy, alkoxy, aryloxy or aralkoxyclasses or from other classes, which provides a compound withgood-to-excellent P2Y₁₂ receptor-binding properties, but which does notyield a compound with undesirable properties like chemical instabilityin a formulation, or one with levels of toxicity that are notwell-tolerated by a treated mammal, or especially, not well-tolerated bya human.

Diastereomers are stereoisomers (isomers of identical constitution butdiffering three-dimensional architecture), which do not bear amirror-image relation to each other.

Pharmaceutically acceptable salts are salts that retain the desiredbiological activity of the parent compound and do not impart undesiredtoxicological effects. Pharmaceutically acceptable salt forms includevarious polymorphs as well as the amorphous form of the different saltsderived from acid or base additions. The acid addition salts can beformed with inorganic or organic acids. Illustrative but not restrictiveexamples of such acids include hydrochloric, hydrobromic, sulfuric,phosphoric, citric, acetic, propionic, benzoic, napthoic, oxalic,succinic, maleic, malic, mesylic, adipic, lactic, tartaric, salicylic,methanesulfonic, 2-hydroxyethanesulfonic, toluenesulfonic,benzenesulfonic, camphorsulfonic, and ethanesulfonic acids. Thepharmaceutically acceptable base addition salts can be formed with metalor organic counterions and include, but are not limited to, alkali metalsalts such as sodium or potassium; alkaline earth metal salts such asmagnesium or calcium; and ammonium or tetraalkyl ammonium salts, i.e.,NX₄ ⁺ (wherein X is C₁₋₄). Other salts such as hydrochlorides,hydrobromides, mesylates, sulfates, acetates, tartrates, etc., are alsocontemplated in this invention. Preferred counterions are monovalentions such as NH₄ ⁺, sodium, lithium, potassium, chloride, bromide,bisulfate, and mesylate, with sodium, potassium, chloride and mesylatebeing most preferred due to ease of manufacture, stability, andphysiological tolerance.

Solvates are addition complexes in which a compound is combined with apharmaceutically acceptable cosolvent in some fixed proportion.Cosolvents include, but are not limited to, water, methanol, ethanol,1-propanol, isopropanol, 1-butanol, isobutanol, tert-butanol, acetone,methyl ethyl ketone, acetonitrile, ethyl acetate, benzene, toulene,xylene(s), ethylene glycol, dichloromethane, 1,2-dichloroethane,N-methylformamide, N,N-dimethylformamide, N-methylacetamide, pyridine,dioxane, and diethyl ether. Hydrates are solvates in which the cosolventis water. It is to be understood that the definition of the compound ofthe present invention encompasses all possible hydrates and solvates, inany proportion, which possess the stated activity.

P2Y₁₂ Receptor Antagonist Compounds

The P2Y₁₂ receptor antagonist compounds useful for preventing ortreating diseases or conditions associated with platelet aggregationand/or platelet activation include compound of general Formula I, and/ortautomers thereof, or a pharmaceutically acceptable salt, solvate, orhydrate thereof:

wherein R_(a) and R_(b) are each independently selected from the groupconsisting of:

-   hydrogen, C₁₋₈ alkyl, C₁₋₈ alkenyl, C₁₋₈ alkynyl, C₃₋₇ cycloalkyl,    C₄₋₇ cycloalkenyl, aralkyl (including saturation and/or unsaturation    in the alkylene portion), aryl, and saturated or unsaturated C₃₋₆    heterocycle; where all rings or chains optionally can bear one or    more desired substituents; or-   R_(a) and R_(b) are taken together to form a ring of 3 to 7 members,    with or without substitution, and with or without heteroatoms in    place of ring carbon atoms;-   R_(c)=H, C₁₋₈ alkyl, C₃₋₇ cycloalkyl, aralkyl, aryl, or heterocycle,    or R(CO)—;-   where R is selected from the group consisting of: C₁₋₈ alkyl, C₁₋₈    alkenyl, C₁₋₈ alkynyl, C₃₋₇ cycloalkyl, C₄₋₇ cycloalkenyl, aryl,    aralkyl, heteroaryl, and saturated or unsaturated C₃₋₆ heterocycle;    where all rings or chains optionally bear one or more desired    substituents;-   G=O, S, or NR_(d), where R_(d) is defined as below;-   R_(d) and R_(d′) are independently selected from the group    consisting of: H, C₁₋₈ alkyl, C₁₋₈ alkenyl, C₁₋₈ alkynyl, C₃₋₇    cycloalkyl, C₄₋₇ cycloalkenyl, C₄₋₁₁ alkylcycloalkyl, C₅₋₁₁    alkylcycloalkenyl, with 1 to 4 carbons in the alkyl portion, aralkyl    (including saturation and/or unsaturation in the alkylene portion),    aryl, heteroaryl, and saturated or unsaturated C₃₋₆ heterocycle; or-   R_(d) and R_(d′) groups are taken together to form a ring of 4 to 7    members, with or without unsaturation and with or without    heteroatoms in place of ring-carbon units; or-   R_(d) or R_(d′) and R_(c) are taken together to form a ring of 4 to    7 members, with or without unsaturation and with or without    heteroatoms in place of ring-carbon units;-   R_(e)=O or absent;-   R_(f)=H, halogen, C₁₋₈ alkyl, C₁₋₈ alkenyl, C₁₋₈ alkynyl, C₃₋₇    cycloalkyl, C₄₋₇ cycloalkenyl, C₄₋₁₁ alkylcycloalkyl, C₅₋₁₁    alkylcycloalkenyl, with 1 to 4 carbons in the alkyl portion, aryl,    aralkyl (including saturation and/or unsaturation in the alkylene    portion), heteroaryl, saturated or unsaturated C₃₋₆ heterocycle,    —OH, C₁₋₆ alkoxy, aryloxy, —SH, C₁₋₆ thioalkyl, thioaryl, —[(CO)OR],    —[(CO)NRR], amino, —N-substituted amino, or N,N-disubstituted amino;    wherein each said substituent on said N-substituted-amino group, or    N,N-disubstituted-amino-group of R_(f) is independently selected    from the group consisting of:-   C₁₋₈ alkyl, C₁₋₈ alkenyl, C₁₋₈ alkynyl, C₃₋₇ cycloalkyl, C₄₋₇    cycloalkenyl, aryl, aralkyl, heteroaryl, C₃₋₆ heterocycle, —[(CO)R]    and —[(CO)—NRR]; wherein each R is independently as defined above;    or-   when R_(f) is —NRR, —[NH(CO)NRR], —[N(C₁₋₈ alkyl)(CO)NRR],    —[N(aryl)(CO)NRR], or [N(aralkyl)(CO)NRR], the R groups of said —NRR    unit (N,N-disubstituted-amino-group) in R_(f) can be taken together    such that a ring of 3 to 7 members is formed, with or without    heteroatoms in place of the ring-carbon units;-   J=N or C, with the proviso that when J=N, then R_(g) is absent;-   when J=C, R_(g) is selected from the group consisting of: —H,    halogen, C₁₋₈ alkyl, C₁₋₈ alkenyl, C₁₋₈ alkynyl, C₃₋₇ cycloalkyl,    C₄₋₇ cycloalkenyl, aralkyl, aryl, —OH, C₁₋₆ alkoxy, aryloxy, —SH,    C₁₋₆ thioalkyl, thioaryl, —[(CO)OR], —[(CO)NRR], and —NRR; wherein    each R is independently as defined above; or-   when R_(g) is —[(CO)NRR] or —NRR, the R groups of said —NRR unit    (N,N-disubstituted-amino-group) in R_(g) are taken together such    that a ring of 3 to 7 members is formed, with or without heteroatoms    in place of the ring-carbon units;-   D is O, NH, N-acyl, N-alkyl, or C;-   A and B are each independently selected from the group consisting    of: C, N, substituted N, O, S, S(O), SO₂, —C₁₋₃ alkylene-, —C₁₋₃    heteroalkylene, wherein each said —C₁₋₃ alkylene-unit of A and B    independently can be saturated or unsaturated, and each carbon of a    —C₁₋₃ alkylene-unit of B independently can be substituted with 0 to    2 fluorine groups, 0 to 1 methyl groups, 0 to 2 —[(CO)OR] groups,    and 0 to 1 —(OR) groups, —CF₂—, —(CO)—; —NH(CO)—, —NR(CO)—,    —(CO)NH—, —(CO)NR—, —NH(CO)NH—, —NH(CS)NH—, —N(NH)NH—, —N(NR)NH—,    —NH(CO)O—, —NH(CS)O—, —O(CO)NH—, —O(CS)NH—, provided that no —S—S—    or —O—O— bonds are formed by combination of the -A- and -B- groups;    or-   A and/or B are absent;-   X=H, —OR, —COOH, —COOR, —SR, —S(O)RL, —S(O₂)RL, —SO₃H, —S(O₂)NRR,    —S(O₂)NR(CO)RL, —NRR, —NR(CO)RL, —N[(CO)RL]₂, —NR(SO₂)RL,    —NR(CO)NR(SO₂)RL, —NR(SO₂)NRR, or —NR(SO₂)NR(CO)RL; wherein L is:-   H, —CF₃, —CF₂CF₃, C₁₋₈ alkyl, C₁₋₈ alkenyl, C₁₋₈ alkynyl, C₃₋₇    cycloalkyl, C₄₋₇ cycloalkenyl, C₄₋₁₁ alkylcycloalkyl, C₅₋₁₁    alkylcycloalkenyl, with 1 to 4 carbons in the alkyl portion,    saturated or unsaturated heteroaryl, aryl, aralkyl (including    saturation and/or unsaturation in the alkylene portion), saturated    or unsaturated C₃₋₆ heterocycle, C₁₋₆ alkoxy, aralkoxy, aryloxy,    N,N-disubstituted-amino, N-substituted-amino, or    unsubstituted-amino; where all rings or chains optionally bear one    or more desired substituents; or-   when L is N-substituted-amino, or N,N-disubstituted-amino, each    substituent of said amino group of L is selected from the group    consisting of: C₁₋₈ alkyl, C₁₋₈ alkenyl, C₁₋₈ alkynyl, C₃₋₇    cycloalkyl, C₄₋₇ cycloalkenyl, aryl, heteroaryl, aralkyl, and C₃₋₆    heterocycle;-   when L is N,N-disubstituted-amino, the two substituents    independently selected from the group above are taken together to    form a ring of 3 to 7 members, wherein said formed ring thereon    bears the remaining features of said selected substituents before    said ring formation; optionally can be made for any one-carbon-unit    within either or both of said C₁₋₃ alkylene units of A and B,    provided that fewer than three said heteroatom-containing-unit for    -one-carbon-unit substitutions on the -A-B- chain are made, no    —S—S—, or —O—O— bonds are formed in the X-A-B- chain by said    substitution or substitutions of a heteroatom-containing-unit for a    -one-carbon-unit on the -A-B- chain, and no said heteroatom    substitution is made such that the said replacement heteroatom    connects directly to the tetrahydrofuran ring shown in Formula I;-   wherein the R groups of a —NRR unit (N,N-disubstituted-amino-group)    in X optionally can be taken together such that a ring of 3 to 7    members is formed, with or without heteroatoms in place of the    ring-carbon units;-   with the proviso that when X=H, then at least one of R_(a) or R_(b)    must be H; or-   X is a group as provided in Formula II:

-   wherein:    -   X₆ is the attachment point to the moiety defined by A-B;    -   the ring defined by X₁-X₆ is taken to mean a ring with or        without unsaturation;    -   X₁-X₆ are independently C, N, O, or S; and    -   when any of X₁-X₅ is C, the carbon atom bears an H when doubly        bonded in an unsaturated ring, or a substituent M, as defined        below; or    -   when any of X₁-X₅ is C, the carbon atom bears two H when singly        bonded in a saturated ring, or one H plus one substituent M, or        two substituents M without H, with the proviso that any such        moiety with one or two M substituents is of sufficient chemical        stability;    -   when any of X₁-X₅ is N in an saturated ring, the nitrogen atom        bears an H or substituents such as alkyl or acyl;    -   any of X₁-X₅ can be absent, with the proviso that at least two        of X₁-X₅ are present, such that the ring described by X₁-X₆        consists of at least three atoms;    -   with the provisos that no two adjacent atoms X₁-X₆ can both be O        or S, and that the ring shown in Formula II contains no more        than four heteroatoms, and that the shown pendant —CO₂R_(h) unit        in Formula II is a substituent on the ring described in Formula        II;    -   p=0, 1, or 2;    -   r=0 or 1;    -   R_(h) is H, a physiologically-relevant cation forming a        carboxylate salt, alkyl, aryl, or aralkyl, with the resultant        moiety C(O)OR_(h) preferably having an adjacent relationship to        the attachment point of A; preferably R_(h) is H or alkyl (such        as ethyl):    -   M is selected from the group consisting of: —H, halogen (such as        F, Cl, Br), —CF₃, C₁₋₈ alkyl, C₁₋₈ alkenyl, C₁₋₈ alkynyl, C₃₋₇        cycloalkyl, C₄₋₇ cycloalkenyl, aryl, aralkyl, heteroaryl,        saturated or unsaturated C₃₋₆ heterocycle, —OH, cyano, saturated        or unsaturated C₁₋₆ alkoxy, aralkoxy, aryloxy, —SH, C₁₋₆        thioalkyl, thioaryl, —[(CO)OR], —[(CO)NRR], amino,        —N-substituted amino, and N,N-disubstituted amino; wherein each        said substituent on said amino of M is independently selected        from the group consisting of: C₁₋₈ alkyl, C₁₋₈ alkenyl, C₁₋₈        alkynyl, C₃₋₇ cycloalkyl, C₄₋₇ cycloalkenyl, aryl, aralkyl,        heteroaryl, C₃₋₆ heterocycle, —[(CO)R], —[(CO)O—(C₁₋₈ alkyl)],        and —[(CO)—NRR]; and

more than one moiety M can be present, either the same or different.

Preferably, the furanosyl moiety in Formula I has the 2′- and3′-oxygen-groups in a cis-orientation relative to one another on thefuranose ring. Further, a furanosyl moiety which supports a 2′,3′-acetalor -ketal group is, preferably, derived from ribose; other furanosederivatives can be used, however. A preferred stereochemical embodimentof this invention includes, but is not limited to(D)-ribose-(2′,3′-acetal or -ketal) compounds of Formula I, such asfound in acetals derived from (−)-adenosine.

In one embodiment of the method, the compound of Formula I is selectedfrom the group consisting of:

4-{2,2-Dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-isophthalicacid (1),5-Amino-2-{2-benzyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-benzoicacid (2),3-{2,2-Dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-isoxazole-5-carboxylicacid (3),4-{2,2-Dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-benzoicacid (4),5-Amino-2-{2,2-dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-N-hydroxy-benzamide(5),5-Amino-2-{2-benzyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-N-hydroxy-benzamide(6),6-{2,2-Dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinamide(7),6-{2,2-Dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinicacid (8),2-{2,2-Dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinicacid (9),5-Chloro-6-{2,2-dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinicacid (10),1-{9-[6-(3-Hydroxy-pyridin-2-yloxymethyl)-2,2-dimethyl-tetrahydro-furo[3,4-d][1,3]dioxol-4-yl]-9H-purin-6-yl}-3-phenyl-urea(11),6-Chloro-2-{2,2-dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-5-fluoro-nicotinicacid (12),2-{2-Cyclohexyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinicacid (13),2-[6-[6-(3-Phenyl-ureido)-purin-9-yl]-2-(2-trifluoromethyl-phenyl)-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy]-nicotinicacid (14),2-{2-(3,4-Dihydro-1H-naphthalenyl)-6-[6-(3-cyclopentyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinicacid (15),2-{2-(4-Acetylamino-phenyl)-6-[6-(3-cyclopentyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinicacid (16),2-{2-Phenyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinicacid (17),2-{2-Biphenyl-3-yl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinicacid (18),2-{2-Naphthalen-2-yl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinicacid (19),2-{2-(2-Bromo-phenyl)-6-[6-(3-ethyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinicacid (20),2-{2-Benzo[b]thiophen-3-yl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinicacid (21),2-{6-[6-(3-Cyclopentyl-ureido)-purin-9-yl]-2-phenethyl-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinicacid (22),2-{6-[6-(3-Cyclopentyl-ureido)-purin-9-yl]-2-phenethyl-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinicacid (23),2-{6-[6-(3-Hexyl-ureido)-purin-9-yl]-2-phenyl-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinicacid (24),2-{2-Biphenyl-4-yl-6-[6-(3-hexyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinicacid (25),2-{6-[6-(3-Ethyl-ureido)-purin-9-yl]-2-phenylethynyl-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinicacid (26),2-{6-[6-(3-Ethyl-ureido)-purin-9-yl]-2-phenethyl-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinicacid (27),2-{6-[6-(3-Cyclopentyl-ureido)-purin-9-yl]-2-p-tolyl-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinicacid (28),2-{2-(2-indanonyl)-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinicacid (29),2-{6-[6-(3-Ethyl-ureido)-purin-9-yl]-2-phenyl-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinicacid (30),2-{2-tert-Butyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinicacid (31),3-({2,2-Dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxole-4-carbonyl}-amino)-benzoicacid (32),2-Benzyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxole-4-carboxylicacid (33),1-{2-Benzyl-6-[6-(3-ethyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxole-4-carbonyl}-pyrrolidine-2-carboxylicacid (34),1-{6-[6-(3-Benzyl-ureido)-purin-9-yl]-2-phenyl-tetrahydro-furo[3,4-d][1,3]dioxole-4-carbonyl}-pyrrolidine-2-carboxylicacid (35),1-{2-Benzyl-6-[6-(3-cyclopentyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxole-4-carbonyl}-pyrrolidine-2-carboxylicacid (36),N-{2-Benzyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxole-4-carbonyl}-methanesulfonamide(37),1-{6-[6-(3-Cyclopentyl-ureido)-purin-9-yl]-2-phenyl-tetrahydro-furo[3,4-d][1,3]dioxole-4-carbonyl}-pyrrolidine-2-carboxylicacid (38),1-{2-Phenyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxole-4-carbonyl}-pyrrolidine-2-carboxylicacid (39),1-{2-Benzo[b]thiophen-3-yl-6-[6-(3-hexyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxole-4-carbonyl}-pyrrolidine-2-carboxylicacid (40),1-{6-[6-(3-Benzyl-ureido)-purin-9-yl]-2-naphthalen-2-yl-tetrahydro-furo[3,4-d][1,3]dioxole-4-carbonyl}-pyrrolidine-2-carboxylicacid (41),1-(2-Benzyl-6-{6-[3-(2-phenyl-cyclopropyl)-ureido]-purin-9-yl}-tetrahydro-furo[3,4-d][1,3]dioxole-4-carbonyl)-pyrrolidine-2-carboxylicacid (42),1-{2-Benzyl-6-[6-(3-hexyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxole-4-carbonyl}-pyrrolidine-2-carboxylicacid (43),1-{2-(2,4-Difluoro-phenyl)-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxole-4-carbonyl}-pyrrolidine-2-carboxylicacid (44),2-({2-Benzyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxole-4-carbonyl}-amino)-3-hydroxy-propionicacid (45),3-{2,2-Dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-yl}-acrylicacid methyl ester (46),3-{2,2-Dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-yl}-propionicacid methyl ester (47),3-(3-{2,2-Dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-yl}-propionylamino)-benzoicacid (48),1-(3-{2,2-Dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-yl}-propionyl)-pyrrolidine-2-carboxylicacid (49), and3-{2,2-Dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-yl}-propionicacid (50). The exemplified compounds named above can be in the formsdepicted below, or can be pharmaceutically-acceptable salts, -hydrates,or -solvates thereof, where chemically appropriate.

In one embodiment of the present invention, when R_(a) and R_(b) are notidentical, the compounds depicted in the following structures fallingunder the definitions of Formulae III-XII represent either one of thetwo possible diastereomers (which arise from the resultant chiral carbonof the acetal) in pure form, or a mixture of the two diastereomers inany proportion. As a practical matter however, the compounds as depictedrepresent the pure forms of the diastereomers. Diastereomers aredistinct compounds, each with potentially different chemical andbiological properties; thus pure forms are preferred as pharmaceuticalagents. In addition, there are generally reasons, including but notlimited to, the ease of chemical synthesis or separation, chemical orbiological stabilility, toxicity, pharmacokinetic or pharmacodynamicproperties in living systems, and the like, to choose between the twopossible isomers. While it is possible to resolve such diastereomericmixtures using chiral chromatographic methods, more preferred is thesynthesis of a single diastereomer.

Depending on the acetal in question, the synthesis of a singlediastereomer can be achieved in several ways. In some cases, onediastereomer can be selectively generated over the other by carrying outthe acetal-forming reaction at a low temperature (such as below 0° C.,for example, from −10 to −30° C.). In other cases, a mixture of twodiastereomers having different acetal stabilities can be subjected toaqueous acidic conditions, which leads to decomposition of theless-stable diastereomer, while leaving the more stable diastereomerintact. In general, the single diastereomer that survives thedecomposition is preferred, since chemical stability is an importantattribute for a pharmaceutical product. These principles are exemplifiedand illustrated in the following compound examples, but as they can bereasonably expanded to related structures; the specific example shouldnot be taken as limiting.

In one embodiment of the present invention, the compound of Formula I isa compound of Formula III:

-   wherein R_(a), R_(b), R_(c), G, R_(d), R_(d)′, R_(e), R_(f), J,    R_(g), and R_(h) are as defined in Formulae I and II;-   A₁ is O or CH₂;-   D is O or CH₂;-   X₁ is selected from the group consisting of: N (nitrogen) and C-M;    and-   M is independently selected from the group consisting of: —H,    halogen, —CF₃, C₁₋₈ alkyl, cyano, C₁₋₈ alkenyl, C₁₋₈ alkynyl, C₃₋₇    cycloalkyl, C₃₋₇ cycloalkenyl, aryl, aralkyl (including saturation    and/or unsaturation in the alkylene portion), heteroaryl, saturated    or unsaturated C₂₋₆ heterocycle, —OH, saturated or unsaturated C₁₋₆    alkoxy, aralkoxy, aryloxy, —SH, C₁₋₆ thioalkyl, thioaryl, —[(CO)OR],    —[(CO)NRR], amino, —N-substituted amino, and N,N-disubstituted    amino; wherein each said substituent on said amino of M is    independently selected from the group consisting of: C₁₋₈ alkyl,    C₃₋₇ cycloalkyl, aryl, aralkyl, heteroaryl, C₂₋₆ heterocycle,    —[(CO)R], —[(CO)O—(C₁₋₈ alkyl)], and —[(CO)—NRR]; and when M is    —[(CO)NRR], —[NH(CO)NRR], —[N(C₁₋₈ alkyl)(CO)NRR],    —[N(aryl)(CO)NRR], or —[N(aralkyl)(CO)NRR], the R groups of any said    —NRR unit (N,N-disubstituted-amino group) in M are optionally taken    together such that a ring of 3 to 7 members is formed, with or    without heteroatoms in place of the ring-carbon units.

Particularly useful compounds of Formula III are where the R_(h)=H oralkyl.

Preferred compounds of Formula III are:

wherein G=A₁=D=O;

R_(a)=R_(c)=R_(d)=R_(f)=R_(g)=R_(h)=H;

R_(d′)=C₁₋₄ alkyl, or C₃₋₆ cycloalkyl;

R_(e) is absent;

X₁=C or N;

R_(b)=phenyl, benzyl, or styryl;

M=H, halogen, C₁₋₄ alkyl, C₁₋₄ alkoxy, CF₃, cyano, or amino.

Some of the preferred compounds falling under the definition of FormulaIII are:

In another embodiment of the present invention, the compound of FormulaI is a compound of Formula IV:

-   wherein R_(a), R_(b), R_(c), G, D, R_(d), R_(d′), R_(e), R_(f), J,    M, R_(g) and R_(h) are as defined in Formulae I and II;-   q₁ and q₂ are 0, 1, or 2;-   the M and —CO₂R_(h) groups are independently and optionally attached    to any carbon of the pyrrolidine ring; and-   when M is attached to a carbon that is bonded to the pyrrolidine    nitrogen atom (alpha position), then M is not a halogen, hydroxyl,    sulfhydryl, or amino group.

Particularly useful groups of compounds are those of Formula IV whereR_(h) is H or alkyl and/or M is H or alkyl.

Preferred compounds of Formula IV are wherein:

q₁ is 1 or 2;

q₂ is 0 or 1;

G=O;

D=O or C;

R_(a)=R_(c)=R_(d)=R_(f)=R_(g)=H;

R_(e) is absent;

R_(h)=H or ethyl;

R_(d′)=C₁₋₄ alkyl, or C₃₋₆ cycloalkyl;

R_(b)=phenyl, benzyl, or styryl;

M=H or C₁₋₄ alkyl.

Some of the preferred compounds falling under the definition of FormulaIV are:

In another embodiment of the present invention, the compound of FormulaI is a compound of Formula V:

-   wherein R_(a), R_(b), R_(c), G, D, R_(d), R_(d′), R_(e), R_(f), J,    M, X₁, R_(g) and R_(h) are as defined in Formulae I and II;-   A₂ is C, O, S, S(O), SO₂, or N, where C can be substituted with H or    alkyl, and N can be substituted with H, alkyl, or acyl; or-   A₂ is absent.

Preferred compounds of Formula V are wherein:

-   G=O;-   D=O or C;-   R_(a)=R_(c)=R_(d)=R_(f)=R_(g)=H;-   R_(e) is absent;-   R_(h)=H or ethyl;-   R_(d′)=C₁₋₄ alkyl, or C₃₋₆ cycloalkyl;-   A₂ is C, O, NH, N-methyl, N-acetyl, or absent;-   X₁=C or N;-   R_(b)=phenyl, benzyl, or styryl; and-   M=H, halogen, C₁₋₄ alkyl, C₁₋₄ alkoxy, CF₃, cyano, or amino.

Some of the preferred compounds falling under the definition of FormulaV are:

In another embodiment of the method, the compound is a compound ofFormula VI:

wherein:

-   R_(a), R_(b), R_(c), G, D, R_(d), R_(d′), R_(e), R_(f), J, M, X₁,    R_(g) and R_(h) are as defined in Formulae I and II;-   A₃ is C, where C can be substituted with H or alkyl; or-   A₃ is absent;-   R_(i) is H or alkyl.

Preferred compounds of Formula VI are wherein:

-   G=D=O;-   R_(a)=R_(c)=R_(d)=R_(e)=R_(f)=R_(g)=R_(h)=R_(i)=H;-   R_(d′)=C₁₋₄ alkyl, or C₃₋₆ cycloalkyl;-   A₃=CH₂, or absent;-   X₁=C or N;-   R_(b)=phenyl, benzyl, or styryl; and-   M=H, halogen, C₁₋₄ alkyl, C₁₋₄ alkoxy, CF₃, cyano, or amino.

Some of the preferred compounds falling under the definition of FormulaVI are:

In another embodiment of the present invention, the compound of FormulaI is a compound of Formula VII:

wherein:

-   R_(a), R_(b), R_(c), G, D, R_(d), R_(d′), R_(e), R_(f), J, R_(g) and    R_(h) are as defined in Formulae I and II;-   q₁ and q₂ are independently 0, 1, or 2;-   A₂ is as previously defined for Formula V, with the proviso that    when q₁ and/or q₂ are 0 and D=O, A₂ is C; or-   A₂ is absent.

Preferred compounds of Formula VII are wherein:

-   G=D=O;-   R_(a)=R_(c)=R_(d)=R_(f)=R_(g)=H;-   R_(e) is absent;-   R_(h)=H or ethyl;-   R_(d′)=C₁₋₄ alkyl, or C₃₋₆ cycloalkyl;-   A₂=CH₂, O, NH, N-methyl, N-acetyl, or absent;-   q₁ and q₂=0 or 1; and-   R_(b)=phenyl, benzyl, or styryl.

Some of the preferred compounds falling under the definition of FormulaVII are:

In another embodiment of the present invention, the compound of FormulaI is a compound of Formula VIII:

wherein:

-   R_(a), R_(b), R_(c), G, D, R_(d), R_(d′), R_(e), R_(f), J, R_(g) and    R_(h) are as defined in Formulae I and II;-   q₃ is 1, 2, or 3; and-   R_(i) is H or alkyl.

Preferred compounds of Formula VIII are wherein:

-   G=D=O;-   R_(a)=R_(c)=R_(d)=R_(f)=R_(g)=H;-   R_(e) is absent;-   R_(h)=H or ethyl;-   R_(i) is H or methyl;-   R_(d′)=C₁₋₄ alkyl, or C₃₋₆ cycloalkyl;-   q₃=1 or 2;-   R_(b)=phenyl, benzyl, or styryl.

Some of the preferred compounds falling under the definition of FormulaVIII are:

In another embodiment of the present invention, the compound of FormulaI is a compound of Formula IX:

wherein:

-   R_(a), R_(b), R_(c), G, D, R_(d), R_(d′), R_(e), R_(f), J, R_(g) and    R_(h) are as defined in Formulae I and II;-   A₄ and A₆ are independently C, N, O, or S, with the proviso that A₄    can be absent;-   G′ is O, or S;-   such that the moiety described by A₄/C(G′)/A₆ is an amide,    thioamide, carbamate, thiocarbamate, urea, thiourea, ketone, or    thioketone;-   q₁ is 0, 1, or 2.

Preferred compounds of Formula IX are wherein:

-   G=G′=O;-   A₄ and A₆ are independently C, N, or O;-   R_(a)=R_(c)=R_(d)=R_(e)=R_(f)=R_(g)=R_(h)=H;-   R_(d′)=C₁₋₄ alkyl, or C₃₋₆ cycloalkyl;-   q₁=1; and-   R_(b)=phenyl, benzyl, or styryl.

Some of the preferred compounds falling under the definition of FormulaIX are:

In another embodiment of the present invention, the compound of FormulaI is a compound of Formula X:

wherein:

-   R_(a), R_(b), R_(c), G, D, R_(d), R_(d′), R_(e), R_(f), J, M, X₁,    R_(g) and R_(h) are as defined in Formulae I and II;-   G′ is O or S;-   A₆ is C, N, O, S, or absent; and-   R_(i) is H or alkyl;-   such that the moiety described by A₆/C(G′)/NR_(i) is an amide,    thioamide, carbamate, thiocarbamate, urea, or thiourea.

Preferred compounds of Formula X are wherein:

-   G=G′=O;-   D=O or C;-   R_(a)=R_(c)=R_(d)=R_(f)=R_(g)=R_(h)=H;-   R_(e) is absent:-   R_(i)=H or methyl;-   R_(d′)=C₁₋₄ alkyl, or C₃₋₆ cycloalkyl;-   A₆=CH₂, O, NH, or absent;-   X₁=C or N;-   R_(b)=phenyl, benzyl, or styryl; and-   M=H, halogen, C₁₋₄ alkyl, C₁₋₄ alkoxy, CF₃, cyano, or amino.

Some of the preferred compounds falling under the definition of FormulaX are:

In another embodiment of the present invention, the compound of FormulaI is a compound of Formula XI:

wherein:

-   R_(a), R_(b), R_(c), G, D, R_(d), R_(d′), R_(e), R_(f), J, M, R_(g)    and R_(h) are as defined in Formulae I and II;-   R_(i) is H or alkyl;-   G′ is O or S, such that the moiety C(G′)-NR_(i) is an amide or    thioamide; and-   X₁ is C or N.

Preferred compounds of Formula XI are wherein:

-   G=G′=O;-   D=O or C;-   R_(a)=R_(c)=R_(d)=R_(f)=R_(g)=R_(h)=H;-   R_(e) is absent;-   R_(i)=H or methyl;-   R_(d′)=C₁₋₄ alkyl, or C₃₋₆ cycloalkyl;-   X₁=C or N;-   R_(b)=phenyl, benzyl, or styryl; and-   M=H, halogen, C₁₋₄ alkyl, C₁₋₄ alkoxy, CF₃, cyano, or amino.

Some of the preferred compounds falling under the definition of FormulaXI are:

In another embodiment of the present invention, the compound of FormulaI is a compound of Formula XII:

wherein:

-   R_(a), R_(b), R_(c), G, D, R_(d), R_(d′), R_(e), R_(f), J, M, R_(g)    and R_(h) are as defined in Formulae I and II;-   X₁, X₂, X₄, X₅, and X₆ are taken to mean a ring with or without    unsaturation and are independently selected from the group    consisting of: N, C, S, or O; and-   X₁-X₆ are taken to mean a ring of from three to five atoms;-   q₁ and q₂ are independently 0, 1, or 2;-   A₂ is C, O, S, S(O), SO₂, or N, or-   A₂ is absent;-   such that when q₁ and q₂=0 and A₂ is absent, the ring described by    X₁/X₂/X₄/X₅/X₆ is directly bonded to the 4′ position of the ribose.

Preferred compounds of Formula XII are wherein:

-   G=O;-   D=O or C;-   R_(a)=R_(c)=R_(d)=R_(f)=R_(g)=R_(h)=H;-   R_(e) is absent;-   q₁ and q₂=0 or 1;-   A₂=CH₂, O, NH, or absent;-   R_(d′)=C₁₋₄ alkyl, or C₃₋₆ cycloalkl;-   R_(b)=phenyl, benzyl, or styryl; and-   M=H.

Some of the preferred compounds falling under the definition of FormulaXII are:

Pharmaceutical Formulations

The present invention additionally provides novel pharmaceuticalformulations comprising a pharmaceutically acceptable carrier andcompounds of Formula I, III-XI, or a pharmaceutically acceptable salt,solvate, or hydrate thereof. Pharmaceutically acceptable carriers can beselected by those skilled in the art using conventional criteria.Pharmaceutically acceptable carriers include, but are not limited to,saline solution, aqueous electrolyte solutions, isotonicy modifiers,water polyethers such as polyethylene glycol, polyvinyls such aspolyvinyl alcohol and povidone, cellulose derivatives such asmethylcellulose and hydroxypropyl methylcellulose, polymers of acrylicacid such as carboxypolymethylene gel, polysaccharides such as dextrans,and glycosaminoglycans such as sodium hyaluronate and salts such assodium chloride and potassium chloride.

The pharmaceutical formulation of the present invention provides anaqueous solution comprising water, suitable ionic or non-ionic tonicitymodifiers, suitable buffering agents, and a compound of Formula I, Ia,Ib, or Ic. In one embodiment, the compound is at 0.005 to 3% w/v, andthe aqueous solution has a tonicity of 200-400 mOsm/kG and a pH of 4-9.

The pharmaceutical formulation can be sterilized by filtering theformulation through a sterilizing grade filter, preferably of a0.22-micron nominal pore size. The pharmaceutical formulation can alsobe sterilized by terminal sterilization using one or more sterilizationtechniques including but not limited to a thermal process, such as anautoclaving process, or a radiation sterilization process, or usingpulsed light to produce a sterile formulation. In one embodiment, thepharmaceutical formulation is a concentrated solution of the activeingredient; the formulation can be serially diluted using appropriateacceptable sterile diluents prior to intravenous administration.

In one embodiment, the tonicity modifier is ionic such as NaCl, forexample, in the amount of 0.5-0.9% w/v, preferably 0.6-0.9% w/v.

In another embodiment, the tonicity modifier is non-ionic, such asmannitol, dextrose, in the amount of at least 2%, or at least 2.5%, orat least 3%, and no more than 7.5%; for example, in the range of 3-5%,preferably 3.5-5%, and more preferably 4.2-5% w/v.

Those skilled in the art will recognize various synthetic methodologiesthat may be employed to prepare non-toxic pharmaceutically acceptablesalts and prodrugs of the compounds.

Methods of Compound Preparation

The compounds of the present invention can be synthesized by thoseskilled in the art using conventional synthesis methodology andwell-known workup and purification procedures. The following list ofreferences, along with references cited therein, disclose generalprocedures employed for the synthesis of a number of intermediates andcompounds related to the present invention: Baraldi, et al., Journal ofMedicinal Chemistry, 39(3): 802-806 (1996); Camaioni, et al., Bioorganic& Medicinal Chemistry, 5(12): 2267-2275 (1997); Zablocki, et al., PCTInternational Publication No. WO01/40243; Zablocki, et al., PCTInternational Publication No. WO01/40246; Mantell, et al., PCTInternational Publication No. WO01/94368; Jacobson, et al., Journal ofMedicinal Chemistry, 38(10): 1720-1735 (1995); Cristalli, et al.,Journal of Medicinal Chemistry, 38(9): 1462-1472 (1995); Secrist, IIIand Talekar, Nucleosides & Nucleotides, 9(4): 619-27 (1990); Secrist,III, U.S. Pat. No. 4,794,174 (1988); Lyga and Secrist, III, Journal ofOrganic Chemistry, 48(12): 1982-1988 (1983); Dixon, et al., PCTInternational Publication No. WO02/096248; Hardern, et al., PCTInternational Publication No. WO01/36438; Guile et al., PCTInternational Publication No. WO00/04021; Lee, et al., Bioorganic &Medicinal Chemistry Letters, 13(6): 1087-1092 (2003); Cox, et. al., U.S.Pat. No. 5,747,496 (1998).

In many cases, commercially-available starting materials can be used forthe synthesis of compounds of this invention. When not availablecommercially, useful starting materials can either be obtained fromstepwise modification of commercially-available compounds andderivatives, or they can be synthesized from simpler precursors usingliterature methods known in the art. In addition, the compounds of thepresent invention can be synthesized using the general methods shown inSchemes 1-12, or variations thereof.

Commercially-available materials include: adenosine, -adenosine,2′,3′-isopropylidineadenosine, 5′-acetyl-2′,3′-isopropylidineadenosine,N⁶-(2-isopentenyl)adenosine, 2-chloroadenosine, 2-amino-6-chloropurineriboside, 6-chloropurine riboside, inosine, 8-bromoguanosine,8-bromoadenosine, 8-azidoadenosine, 8-azaguanine, 8-azaadenine,protected ribonic acid lactone derivatives and protected furanosederivatives. Other appropriate intermediates can be purchased fromcommercial sources and used as starting materials for compounds of thepresent invention, or can be synthesized as described in the chemicalliterature.

As disclosed above, commercially available compounds, or theirderivatives can be employed as starting materials for the methods ofSchemes 1-12.

Scheme 1. Preparation of 5′-modified ethers by nucleophilic aromaticsubstitution.

Scheme 1, for example, discloses a useful method for the synthesis of5′-aryl- or 5′-heteroaryl-ether derivatives by substitution of anappropriately functionalized adenosine analogue or 8-azapurinederivative for a halogen on an appropriately-substituted halogenatedaromatic compound or a related heteroaromatic derivative. Groups notdefined in Scheme 1 are defined as in Formula I. Preferred substituentsat M of the aromatic-/heteroaromatic-group in Scheme 1 are hydrogen, orhalogen, or groups containing carboxylic acid derivatives such as:—CO₂R₃; but they can also be halogen, or esters or amides ofalkylcarboxylic acids, arylcarboxylic acids, —O-(alkylcarboxylic acids),—NR-(alkylcarboxylic acids), and the like. When M is halogen in Scheme1, preferred halogens are chloro and fluoro.

Scheme 2. Preparation of 5′-modified ethers by Mitsunobu coupling ofphenols.

5′-Substituted aryl derivatives can also be prepared via Mitsunobucoupling of phenols (Mitsunobu, Synthesis 1-28 (1981); Brown, et al., J.Med. Chem. 37 (5), 674-88 (1994); Santosh and Balasubramanian, SyntheticCommunications, 24(8), 1049-62 (1994)) to derivatives of adenosine,8-azaadenosine, guanosine, 8-azaguanosine, etc., as provided in Scheme2. Groups in Scheme 2 are defined as in Formula I. Some preferredsubstituents at M of the aromatic/heteroaromatic-group in Scheme 2independently can be hydrogen, halogen, alkyl, alkoxy, aryl or groupscontaining carboxylic acid derivatives such as: —CO₂R₃; but esters oramides of alkylcarboxylic acids, arylcarboxylic acids,—O-(alkylcarboxylic acids), —NR-(alkylcarboxylic acids), and the likeare also included. When M is halogen in Scheme 2, preferred halogens arechloro and fluoro.

Scheme 3. Preparation of 5′-modified isoxazole ethers by Mitsunobucoupling of isoxazole derivatives.

Alternatively, the Mitsunobu coupling can be carried out usinghydroxyisoxazoles as provided in Scheme 3. Groups in Scheme 3 aredefined as in Formula I.

Some examples of preferred substituents at M of the isoxazolederivatives of Scheme 3 independently include hydrogen, alkoxy, orhalogen, or groups containing carboxylic acid derivatives such as:—CO₂R₃; but esters or amides of alkylcarboxylic acids, arylcarboxylicacids, —O-(alkylcarboxylic acids), —NR-(alkylcarboxylic acids), and thelike are also included. When M is halogen in Scheme 3, preferredhalogens are chloro and fluoro.

When the products of any of Schemes 1, 2 or 3 comprise esters, saidesters can be useful in the invention. Said ester derivatives can bepurified by methodologies well-known in the art, such as by normalphase, or reverse phase chromatography, or in suitable circumstances,using crystallization techniques. Alternatively, said ester derivativescan be used in the synthesis of other derivatives such as amides,hydroxamic acids, and different alkyl esters by well-known methods inthe art.

Optionally, said esters can be hydrolysed under basic conditions, orcleaved using other methods known in the art, which cleave estersselectively in the presence of ketals or acetals to provide acid salts.These salts are also useful in the invention.

If desired, said acid salts can be converted into acids upon mild acidtreatment. Workup by common techniques, and purification by methods wellknown in the art, including purification by crystallization orchromatography can be used to give the purified acids. Said acids canalso be converted into other useful derivatives such as amides,hydroxamic acids, aryl esters, etc., by methods known to those skilledin the art of chemical synthesis. These acid derivatives are also usefulin the invention, and are also purified using well-known methods such ascrystallization or chromatography.

Scheme 4. Transformation of 2′, 3′-acetals and -ketals into new acetaland ketal derivatives using solid phase techniques.

Diversity using common intermediates can be introduced into the2′,3′-acetal or -ketal position of compounds encompassed in Formula I,as well as at the N⁶-position of compounds of Formula I usingsolid-phase synthetic methods. Schemes 4 and 5 exemplifytransketalization procedures using polymer-bound relatives of compoundsof Formula I. These methods can be used to transform one class of2′,3′-ketal or -acetal into other useful 2′,3′-acetal or -ketalderivatives of adenosine, guanosine, 8-azaadenosine, etc. Thesepolymer-bound approaches are very useful, for after the desired reactionis complete, excess reagents can be washed away using one to severalsolvent washes. The desired material remains attached to the resin, in apre-purified form, until it is cleaved from the solid phase using theappropriate conditions. Final purification of the desired product isthen accomplished by conventional techniques like chromatography orcrystallization.

Scheme 5. Transformation of 6-chloro derivatives into 6-urea, 6-thioureaand 6-guanidine derivatives; subsequent conversion (when desired) of 2′,3′-acetals and -ketals into new acetal and ketal derivatives using solidphase techniques.

Scheme 5 shows another variation of compound preparation employingresin-bound materials of Formula I. It outlines a solid-phase procedurethat is useful for the introduction of functionality at the 6-position,as well as for transketalization at the 2′,3′-position, if desired. Itshould be noted that the chemistry procedures used in these solid phaseapproaches are similar to methods known and used for solution-phasechemical transformations involving the synthesis of adenosine,guanosine, 8-azaadenosine, etc., derivatives. The primary differencesare the necessity of attachment of a starting material to a resin, thesimplicity of resin-based purification techniques (filtration andwashing by compatible solvents) compared to solution-phase techniques(chromatography, crystallization, etc.), and the requirement forcleavage of a compound of the invention, or an intermediate useful forthe synthesis of a compound of the invention from a resin prior to finalpurification and/or use of a compound so cleaved.

In Scheme 5, an early intermediate, such as a6-chloroadenosine-2′,3′-ketal or -acetal derivative, or a6-chloro-8-aza-adenosine-2′,3′-ketal or -acetal derivative is attachedto a resin such as polystyrene resin via a β-thioethanol linker (e.g.,hydroxyethylsulfanylmethyl polystyrene; HESM polystyrene resin;Garcia-Echeverria, Tetrahedron Lett., 38, 8933-7 (1997)). After workupby filtration and rinsing with a useful solvent like dimethylformamide(DMF), the resin bound material is treated with a primary amine, ammoniaor a hydroxylamine derivative to introduce an amino group viadisplacement of the 6-chloride. As with solution phase techniques,subsequent introduction of an ureido-, thioureido-, or guanidino-groupat N⁶ can be made in one step using an excess of an appropriateisocyanate, isothiocyanate, carbodiimide, carbamoyl chloride, or2-alkyl-2-thiopseudourea; or using a chemical equivalent of suchmaterials. Alternatively, a two-step approach can be used to introduce agroup at N⁶, which comprises treatment of an appropriate 6-aminoderivative, synthesized as in Scheme 5, with a solution of a smallexcess of phosgene or thiophosgene and a tertiary amine such asdiisopropylethylamine in a suitable solvent such as dichloromethane ortoluene at a temperature which allows reaction, followed by treatmentwith an excess of a primary or secondary amine to give a resin-boundurea or thiourea after workup. If desired, transketalization can beperformed on a bound substrate of Scheme 5 in a manner similar to thatshown in Scheme 4. Or, if a desired acetal or ketal moiety of a6-chloroadenosine, 6-chloroguanosine, 6-chloro-8-azaadenosine, etc.,derivative is used to begin with, then no transketalization isnecessary. Cleavage from the β-thioethanol linker of the solid phase, asshown for the HESM polystyrene resin in Scheme 5 is performed in twosteps: oxidation of the thioether-linker using an oxidizing agent likem-chloroperbenzoic acid in a solvent such as dichloromethane gives asulfone-linker, and cleavage of the β-ether moiety from the oxidizedlinker occurs upon treatment with a strong base like DBU in a solventlike dichloromethane and yields a compound which can be purified bytechniques known in the art. Preferred oxidizing agents include peracidslike m-chloroperbenzoic acid (MCPBA) and peracetic acid, but otheroxidizing agents like hydrogen peroxide, permanganate salts, orpersulfate salts can be used to oxidize a thioether to a sulfone.Preferred elimination conditions include DBU in dichloromethane, and 10%ammonium hydroxide in trifluoroethanol.

Following cleavage from the solid phase, a compound formed using theprocedures in Scheme 5 can be used in the present invention, or can befurther modified by well-known methods for functional-grouptransformations to generate new compounds which are also useful for thisinvention.

Preferred aldehydes, aldehyde acetals and ketone ketals useful in thetransketalization methods shown in Schemes 4 and 5 comprise the belowsaid carbonyl compounds and/or derivatives of: benzaldehyde,biphenyl-3-carboxaldehyde, biphenyl-4-carboxaldehyde,biphenyl-4-yl-acetaldehyde, 2-bromobenzaldehyde,benzo[b]thiophene-3-carbaldehyde, cyclohexanecarbaldehyde,cyclopentanecarbaldehyde, 2,5-dimethylbenzaldehyde,2,6-difluorobenzaldehyde, 2-fluorobenzaldehyde,naphthalene-2-carbaldehyde, phenyl acetaldehyde, phenyl propynal,3-phenyl propenal, 3-phenyl propionaldehyde, 2-trifluoromethylbenzaldehyde, cyclohexanone, cyclopentanone, 4-ethyl cyclohexanone,3,4-dihydro-1H-naphthalen-2-one, and indan-2-one. Useful derivatives fortransketalization of the above ketones comprise ketals like dimethoxy-or diethoxy-ketals, etc.

In addition to the introduction of amines on the C⁶-position using solidphase techniques as provided in Scheme 5, diversity can also beintroduced at the 6-position of an adenosine, 8-azaadenosine, guanosine,etc., analog via the intermediacy of a 6-halogenated-purine derivativeusing solution phase methods. Scheme 6 exemplifies the preparation of5′-isoxazole ethers, introducing ammonia, various amines orhydroxylamine derivatives at the 6-position of the purine/8-azapurinering by displacment of a chloride leaving group (a 6-chloride is shownin Schemes 5 and 6, but the leaving group at C⁶ could also be anothertype, useful for such a transformation, e.g., a 6-bromide or 6-mesylatemoiety) by such materials. Amines and amine-like compounds useful fordisplacement of a 6-halogen intermediate as contemplated for theseschemes comprise ammonia, methylamine and other N-alkyl amines;N-aralkylamines; N-cyclopropylamine and other N-cycloalkylamines;anilines; ethers and other O-derivatives of hydroxylamine;aminopyridines and other heteroaromatic amines; heterocyclic compoundshaving a pendant —NHR_(c)-group; and N-alkyl amines which have one ormore heteroatom units like O, NR, and/or S substituted for carbon unitsin the alkyl chain. Such N⁶-products can be further transformed intoureas, thioureas, or guanidines by literature methods or by methodsdisclosed for such transformations in Schemes 5 and 6. The materials canbe purified by methods typically used in the literature, such as bychromatography or, in certain cases, by crystallization Preferredsubstituents at N⁶ are ureas.

Scheme 6. Solution phase synthesis of 5′-heteroaromaticether-N⁶-derivatives by Mitsunobu reaction, 6-chloride displacement, andN⁶-amine derivatization.

When M of an isoxazole derivative as provided in Scheme 6 contains anester group [e.g., —C(CO)O-(alkyl), —C(CO)O-(aryl),—(CH₂)_(m)C(CO)O-(alkyl), —O(CH₂)_(m)C(CO)O-(aryl), etc., where “m”defines a carbon chain length of a compound of Formula I], said estercan be used in the present invention, or it can be converted into anacid using a method which is compatible with an acetal or ketal moietyand a desired group at N⁶. For example, an ester of Scheme 6 can behydrolysed at room temperature (RT) in several hours using an excess ofaqueous 2M lithium hydroxide solution dissolved in dioxane and/ormethanol to give a carboxylate salt. Purification of said salt or acorresponding acid of said salt can be accomplished as previouslydisclosed. These acids and acid derivatives are also useful in thisinvention. Scheme 6 is exemplified using ammonia and primary amines(including hydroxylamine derivatives) as choices for nucleophiles in thedisplacement of a leaving group (like chloride) at C⁶. Furthermodification of the N⁶-group of a compound of Scheme 6 with a—[(CG)NR_(d)R_(d)′]-group yields a compound useful in the invention.Scheme 7. Synthesis of 2′,3′-O-Isopropylideneadenosine-5′-carboxylicacid, related acetals and ketals, and their use as intermediates for thesynthesis of compounds of this invention.

2′,3′-O-Isopropylideneadenosine-5′-carboxylic acid and related acetalsand ketals are useful intermediates for the preparation of variousamides and sulfonamide derivatives. An intermediate of this type can beused in both solution phase and solid phase synthetic schemes to preparecompounds with various substituents at the 5′-, 2′,3′-acetal/ketal-, and6-positions- of adenosines or related purine derivatives as provided inScheme 7. The methods shown in Scheme 7 are also useful for thepreparation of substituted purine derivatives, and/or 8-azapurinederivatives.

Compounds containing the adenosine 5′-carboxylic acid unit shown inScheme 7, or related N⁶-substituted-adenosine-5′-carboxylic acids and8-azaadenosine-5′carboxylic acids can also be used for the synthesis ofesters or other materials useful in this invention using literaturemethods. In addition, the N⁶-amine of such an acid can be transformedinto a urea, thiourea, or guanidine, or into a protected group such as abenzamide, and the acid moiety which is at, or linked to, the5′-position of the furanose derivative can be coupled to a solid phaseresin such as a 4-sulfamylbutyryl resin, hydroxymethyl resin, or apegylated-hydroxy resin, etc., using techniques well-known in thepeptide literature and/or solid phase organic synthesis literature togive a resin-bound material. The benefits of solid-phase chemistry canthen be gained for modification of a resin-bound material bytransketalization techniques similar to those shown in Schemes 4 and 5.If a protecting group is used on the N⁶-position of such an acid, it canbe removed subsequent to a transketalization procedure, and a N⁶-amineso formed can be converted into a urea, thiourea or guanidine by methodsdisclosed herein, or by methods in the literature. Cleavage from thesolid support using known methods then yields a compound of theinvention which can be purified, if needed, by employing commonly-usedtechniques.

Scheme 8. N⁶-modification of adenosine derivatives, followed byoxidation to a 5′-carboxylic acid, and 5′amide formation.

A variation of the method disclosed in Scheme 7, useful for theintroduction of a group at N⁶ prior to oxidation of the 5′-position ofan adenosine derivative or an 8-azaadenosine derivative, is shown inScheme 8. Although Scheme 8 shows an adenine unit, it will be understoodby chemistry practioners that the methods of Scheme 8 are generallyapplicable to substituted members of the adenine family and also to the8-azaadenines.

Preferred amines and amine derivatives for the 5′-amide-formingreactions shown in Schemes 7 and 8 are: trifluoromethanesulfonamide,methanesulfonamide, serine, glycine, proline, anthranilic acid and itsregioisomers, and methyl anthanilate and its regioisomers.

Scheme 9. Amino acid amides of 5′-adenosinecarboxylic acid derivativesand 8-azaadenosine-5′-carboxylic acid derivatives using solid phasetechniques.

Amide derivatives of 5′-carboxylic acids (e.g., those shown in Scheme 7and Scheme 8) or amide derivatives of acid moieties linked at the5′-position of Formula I can also include amides derived from aminoacids, peptides, aminoalcohols and the like. A convenient way ofattaching naturally-occurring, as well as synthetically-derived aminoacids and peptides, or derivatives, to a 5′-carboxylic acid or relatedhomologue is exemplified in Scheme 9 using the amino acid, proline.

An example of the method is shown in Scheme 9 utilizing a resin-linkercombination like the polystyrene/HESM as a solid phase. Other solidphase/linkers known in the art can also be used in this method.Attachment of a group, such as an amino acid, (e.g., proline, as shown)or a series of amino acids, or a peptide, by well-known methods in theart of solid phase synthesis yields a resin-linked amine. Said amine canthen be reacted with an adenosine 5′-carboxylic acid, or another type ofderivative useful for making compounds of this invention, to yield acoupled product. The said coupled product, if it bears an amine at the6-position, can then be treated with one of the various reagentsdescribed previously, or with a reagent known in the literature, toyield a urea, thiourea or guanidine at the adenosine 6-position.Alternatively, if a 5′-carboxylic acid derivative used in a coupling toa solid phase has a urea, etc., already installed at the 6-position,then the latter said modification at N⁶ need not be performed. Ifdesired, a coupled 5′-amide/N⁶-derivatized product can be converted intoa variety of different acetals or ketals using solid phase methods suchas described for Schemes 4 and 5. When a synthesis is complete, cleavageof a compound of the invention from a solid phase can be performed by avariety of methods known in the art; such cleavage conditions dependingupon the type of linker used. Cleavage methodologies useful for cleavingpeptide- or amino acid derivatives of 5′-linked-adenosine compoundscomprise the linker oxidation/elimination procedures given in Schemes 5and 9; treatment with a hydroxide source, such as lithium hydroxide,using conditions as for the ester hydrolysis described in Scheme 6; andhydrolysis using potassium trimethylsilanolate, as described in Scheme4; as well as others known in the art (including aminolysis to formamides). If desired, a compound useful in this invention can be obtainedin purified form by cleaving it from a resin and purifying it asdescribed previously.

Scheme 10. Synthesis of 5′-amines and 5′-amine derivatives ofnucleosides and 8-azanucleosides.

In another embodiment, an amino group can be installed at the5′-position of an adenosine or 8-azaadenosine analogue, or on the chainof a 5′-homologue of such a material. This amine can be utilized to formamide-, sulfonamide-, and other derivatives. Scheme 10 illustrates how asulfonylurea can be synthesized at the 5′-position using a 5′-amine, orat related positions on homologous amine derivatives. In addition, anamine introduced at the 5′-position or on the 5′-chain of a homologue isalso useful for the synthesis of amides, ureas, sulfonamides and otheramine derivatives using methods known in the art for such processes.

Scheme 11. Homologation of adenosine derivatives, including stepsinvolving oxidation, Wittig or Horner-Emmons Reactions, reduction andcoupling with amines.

In still another embodiment, the 5′-position of the nucleosidederivative or 8-azanucleoside derivative is homologated with one or morecarbon atoms, affording compounds with different distances between theatoms of the tetrahydrofuran ring and the homologated group. Scheme 11illustrates the preparation of a a class of homologated adenosineanalogs which are useful for the invention. Or, if desired, such ahomologue can subsequently be coupled with an amino acid to give othercompounds useful for this invention. In Scheme 11, proline is used toexemplify the amide coupling, but other amines or amino acid derivativescan be employed. In Scheme 12, the reduction step of Scheme 11 isomitted, which generates unsaturated homologues useful in thisinvention.

Scheme 12. Preparation of 5′-substituted ethers.

Scheme 12, for example, discloses a useful method for the synthesis ofaryl- or heteroaryl-nucleoside ethers from purine or 8-azapurinecarbocyclic nucleoside acetals by substitution of an appropriatelyfunctionalized adenosine analogue or 8-azapurine derivative with ahalogenated on an appropriately-substituted halogenated aryl, alkyl, oralkylaryl compound or a related heteroaromatic derivative. Preferredsubstituents at M of the aromatic/heteroaromatic-group in Scheme 12 areindependently hydrogen, or halogen, or groups containing carboxylic acidderivatives such as: —CO₂R₃; but they can also be halogen, or esters oramides of alkylcarboxylic acids, arylcarboxylic acids,—O-(alkylcarboxylic acids), —NR-(alkylcarboxylic acids), and the like.When M is halogen in Scheme 1, preferred halogens are chloro and fluoro.

The method disclosed in Scheme 12 can also be extended to a wide rangeof alkyl or aralkyl halides, making it a useful method for the generalpreparation of 5′ substituted ethers.

Scheme 13. Preparation of 2, 3-acetals and -ketals.

Diversity using common intermediates can be introduced into the2,3-acetal or -ketal position (Scheme 14) of compounds encompassed inFormula I, as well as at the N⁶-position of compounds of Formula I.Final purification of the desired product is then accomplished byconventional techniques like chromatography or crystallization.

Preferred aldehydes, aldehyde acetals and ketone ketals useful in thetransketalization methods shown in Schemes 2 comprise the below saidcarbonyl compounds and/or derivatives of: benzaldehyde,biphenyl-3-carboxaldehyde, biphenyl-4-carboxaldehyde,biphenyl-4-yl-acetaldehyde, 2-bromobenzaldehyde,benzo[b]thiophene-3-carbaldehyde, cyclohexanecarbaldehyde,cyclopentanecarbaldehyde, 2,5-dimethylbenzaldehyde,2,6-difluorobenzaldehyde, 2-fluorobenzaldehyde,naphthalene-2-carbaldehyde, phenyl acetaldehyde, phenyl propynal,3-phenyl propenal, 3-phenyl propionaldehyde, 2-trifluoromethylbenzaldehyde, cyclohexanone, cyclopentanone, 4-ethyl cyclohexanone,3,4-dihydro-1H-naphthalen-2-one, and indan-2-one.

Those having skill in the art will recognize that the starting materialscan be varied and additional steps employed to produce compoundsencompassed by the present invention, as shown in the above schemes andas demonstrated by the examples which follow. In some cases, protectionof certain reactive functionalities can be necessary to achieve some ofthe above transformations. In general, the need for such protectinggroups as well as the conditions necessary to attach and remove suchgroups will be apparent to those skilled in the art.

Use of P2Y₁₂ Receptor Antagonist Compounds

This invention provides a method of preventing or treating diseases orconditions associated with platelet aggregation and/or plateletactivation. This invention also provides a method for solving treatmentproblems or limited treatment options caused by the aggregation ofplatelets or by the irreversible inhibition of platelet aggregation.

This invention provides methods of preventing or treating thrombosis andrelated disorders, such as venous thrombosis, established peripheralarterial disease, thrombophlebitis, arterial embolism, coronary andcerebral arterial thrombosis, unstable angina, myocardial infarction,stroke, cerebral embolism, renal embolism, pulmonary embolism and otherembolism- or thrombosis-related afflictions produced by but not limitedto procedural or surgical interventions. This invention further providesmethods for the prevention of embolism or thrombosis during percutaneouscoronary interventions, placement of coronary stents, coronaryangioplasty, coronary endarectomy, carotid endarectomy, or due toplatelet-aggregation complications related to atherosclerosis,inflammation, exposure of blood to artificial devices, drug effects.

This invention further provides methods of inhibiting plateletaggregation in blood and blood products comprising platelets, such asstored blood.

The method comprises administering to a subject or blood and bloodproducts a composition comprising an effective amount of P2Y₁₂ receptorantagonist compound, wherein said amount is effective to bind the P2Y₁₂receptors on platelets and inhibit platelet aggregation, preferably in areversible manner.

The invention further provides useful methods of treating patients toinhibit platelet aggregation in a reversible manner, especially inpatients that are subject to a procedure such as percutaneous coronaryinterventions, stent placement, balloon angioplasty, coronaryatherectomy, coronary endarterectomy, carotid endarterectomy,thrombolytic theraphy, coronary or other vascular graft surgery,dialysis, etc. In those patients, it is important that plateletaggregation inhibition can be rapidly reversed (within hours for oraladministration and within minutes for intravenous administration) ifnecessary. The method comprises the steps of: (a) providing a patient inneed of rapid reversal of platelet aggregation inhibition; (b)administering a therapeutically effective amount of a compound ofFormula I, Ia, or Ic to the patient; (c) submitting the patient to aprocedure selected from the group consisting of: percutaneous coronaryinterventions, stent placement, balloon angioplasty, coronaryatherectomy, coronary endarterectomy, carotid endarterectomy,thrombolytic theraphy, coronary or other vascular graft surgery, anddialysis, (d) discontinuing the administering of said compound to thepatient; and (e) allowing the amount of said compound in the patient'sblood to reduce to below an therapeutically effective amount. In step(b), the administration of the compound can be either continuous orintermittent as long as it provides a therapeutically effective amountof the compound in the patient's blood. The amount of the compound inthe patient's blood is monitored.

The compounds of general Formulae I, III-XII are antagonists of theeffect of ADP on its platelet membrane receptor, the P2Y₁₂ receptor. Thecompounds of general Formula I are useful in therapy, in particular inthe prevention or treatment of platelet aggregation. The compoundsprovide efficacy as antithrombotic agents by their ability to block ADPfrom acting at its platelet receptor site and thus prevent plateletaggregation. The compounds provide a more efficacious antithromboticeffect than aspirin, but with less profound effects on bleeding thanantagonists of the fibrinogen receptor.

The P2Y₁₂ receptor antagonists of this invention, in contrast withcurrently available marketed products clopidogrel (Plavix®) andticlopidine (Ticlid®), bind to the P2Y₁₂ receptor in a reversiblefashion and therefore, the effects of the treatment with compoundsdescribed in this invention are reversed by the simple discontinuationof the treatment, restoring the hemostatic functionality of the plateletas necessary. Since platelets are non-nucleated cell particles that lackthe ability to synthesize new proteins, treatment of subjects withirreversible P2Y₁₂ antagonists results in the impairment of plateletfunction that lasts for the lifespan of the platelet (approximately 8 to10 days). The use of irreversible P2Y₁₂ antagonists such as clopidogrelhas been associated with increases in blood loss, transfusionrequirements and rate of reoperation after cardiac surgery (Kapetanakis,et al., Eur Heart J. 26: 576-83, 2005). To avoid these complications,subjects undergoing elective surgeries are required to discontinue thetreatment with irreversible antagonists for at least five days prior tothe surgery, which increases the risk of a thrombotic event during thisperiod. Therefore, the compounds described in this invention representan advantage over the currently marketed compounds.

The ADP-induced platelet aggregation is mediated by the simultaneousactivation of both P2Y₁₂ and P2Y₁ receptors, thus the combinedadministration of the Formula I compounds with antagonists of plateletP2Y₁ receptors can provide a more efficacious antithrombotic effect atconcentrations of each antagonist that are below the effectiveconcentrations to block each receptor subtype in other systems,resulting in a decrease of the potential manifestation of adverseeffects. In addition, these compounds can be used in conjunction withlower doses of other platelet aggregation inhibitors, which work bydifferent mechanisms, to reduce the possible side effects of saidagents.

The compounds of the present invention are useful as anti-thromboticagents, and are thus useful in the treatment or prevention of unstableangina, coronary angioplasty (PTCA) and myocardial infarction.

The compounds of the present invention are useful in the treatment orprevention of primary arterial thrombotic complications ofatherosclerosis such as thrombotic stroke, peripheral vascular disease,and myocardial infarction without thrombolysis.

The compounds of the invention are useful for the treatment orprevention of arterial thrombotic complications due to interventions inatherosclerotic disease such as angioplasty, endarterectomy, stentplacement, coronary and other vascular graft surgery.

The compounds of the invention are useful for the treatment orprevention of thrombotic complications of surgical or mechanical damagesuch as tissue salvage following surgical or accidental trauma,reconstructive surgery including skin flaps, and “reductive” surgerysuch as breast reduction.

The compounds of the present invention are useful for the prevention ofmechanically-induced platelet activation in vivo, for example, caused bycardiopulmonary bypass, which results in temporary platelet dysfunction(prevention of microthromboembolism). The compounds of the presentinvention are useful for prevention of mechanically-induced plateletactivation in vitro. For example, the compounds are useful in thepreservation of blood products, e.g. platelet concentrates, preventionof shunt occlusion such as renal dialysis and plasmapheresis, andthrombosis secondary to vascular damage/inflammation such as vasculitis,arteritis, glomerulonephritis and organ graft rejection.

The compounds of the present invention are useful in disorders with adiffuse thrombotic/platelet consumption component such as disseminatedintravascular coagulation, thrombotic thrombocytopenic purpura,hemolytic uremic syndrome, heparin-induced thrombocytopenia andpre-eclampsia/eclampsia.

The compounds of the invention are useful for the treatment orprevention of venous thrombosis such as deep vein thrombosis,veno-occlusive disease, hematological conditions such as thrombocythemiaand polycythemia, and migraine.

The compounds of the present invention are useful in treating a mammalto alleviate the pathological effects of atherosclerosis andarteriosclerosis, acute MI, chronic stable angina, unstable angina,transient ischemic attacks and strokes, peripheral vascular disease,arterial thrombosis, preeclampsia, embolism, restenosis or abruptclosure following angioplasty, carotid endarterectomy, and anastomosisof vascular grafts.

The compounds of the present invention are useful in treating chronic oracute states of hyper-aggregability, such as disseminated intravascularcoagulation (DIC), septicemia, surgical or infectious shock,post-operative and post-partum trauma, cardiopulmonary bypass surgery,incompatible blood transfusion, abruptio placenta, thromboticthrombocytopenic purpura (TTP), snake venom and immune diseases, arelikely to be responsive to such treatment.

The compounds of the present invention are useful in treating diseasesor conditions associated with platelet activation and/or aggregationproduced by the contact of blood with an artificial device. In oneembodiment, the artificial device is a paracorporeal artificial lung andan extracorporeal membrane oxigenation device. In another embodiment,the artificial device is an internal implantable artificial heart. Inanother embodiment, the artificial device is an apheresis instrumentused to remove or isolate a specific component of the blood, andreturning the remaining blood components to the donor. In yet anotherembodiment, the artificial device is a hemodialysis instrument.

The compounds of the present invention are useful in vitro to inhibitthe aggregation of platelets in blood and blood products, e.g. forstorage, or for ex vivo manipulations such as in diagnostic or researchuse. In such applications, the compounds are administered to the bloodor blood product.

Finally, if the compounds of the present invention have sufficientbinding affinity and bear a fluorescent moiety, they are useful asbiochemical probes for the P2Y₁₂ receptor.

In a preferred embodiment, the compounds are used in the treatment ofunstable angina, coronary angioplasty and myocardial infarction.

In another preferred embodiment, the compounds are useful as adjunctivetherapy in the prevention or treatment of thrombotic disorders, such ascoronary arterial thrombosis during the management of unstable angina,coronary angioplasty and acute myocardial infarction, i.e.perithrombolysis. The compounds are administered in combination withother antiplatelet and/or anticoagulant drugs such as heparin, aspirin,GP IIb/IIIa antagonists, or thrombin inhibitors.

This invention provides a method of inhibiting platelet aggregation andclot formation in a mammal, especially a human, which comprisesadministering to the subject a compound of Formula (I) and apharmaceutically acceptable carrier.

This invention further provides a method for inhibiting the reocclusionof an artery or vein following fibrinolytic therapy, which comprisesadministering to a subject a compound of Formula (I) and a fibrinolyticagent. When used in the context of this invention, the term fibrinolyticagent is intended to mean any compound, whether a natural or syntheticproduct, which directly or indirectly causes the lysis of a fibrin clot.Plasminogen activators are a well known group of fibrinolytic agents.Useful plasminogen activators include, for example, anistreplase,urokinase (UK), pro-urokinase (pUK), streptokinase (SK), tissueplasminogen activator (tPA) and mutants, or variants thereof, whichretain plasminogen activator activity, such as variants which have beenchemically modified or in which one or more amino acids have been added,deleted or substituted or in which one or more functional domains havebeen added, deleted or altered such as by combining the active site ofone plasminogen activator or fibrin binding domain of anotherplasminogen activator or fibrin binding molecule.

Extracorporeal circulation is routinely used for cardiovascular surgeryin order to oxygenate blood. Platelets adhere to surfaces of theextracorporeal circuit. Platelets released from artificial surfaces showimpaired hemostatic function. Compounds of the invention can beadministered to prevent adhesion.

Other applications of these compounds include prevention of plateletthrombosis, thromboembolism and reocclusion during and afterthrombolytic therapy and prevention of platelet thrombosis,thromboembolism and reocclusion after angioplasty of coronary and otherarteries and after coronary artery bypass procedures.

The active compounds can be administered systemically to target sites ina subject in need such that the extracellular concentration of a P2Y₁₂antagonist is elevated to block the binding of ADP to P2Y₁₂ receptor,thus inhibit the platelet aggregation. The term systemic as used hereinincludes subcutaneous injection, intravenous, intramuscular,intrasternal injection, intravitreal injection, infusion, inhalation,transdermal administration, oral administration, rectal administrationand intra-operative instillation.

For systemic administration such as injection and infusion, thepharmaceutical formulation is prepared in a sterile medium. The activeingredient, depending on the vehicle and concentration used, can eitherbe suspended or dissolved in the vehicle. Adjuvants such as localanesthetics, preservatives and buffering agents can also be dissolved inthe vehicle. The sterile injectable preparation can be a sterileinjectable solution or suspension in a non-toxic acceptable diligent orsolvent. Among the acceptable vehicles and solvents that can be employedare sterile water, saline solution, or Ringer's solution.

Another method of systemic administration of the active compoundinvolves oral administration, in which pharmaceutical compositionscontaining active compounds are in the form of tablets, lozenges,aqueous or oily suspensions, viscous gels, chewable gums, dispersiblepowders or granules, emulsion, hard or soft capsules, or syrups orelixirs.

For oral use, an aqueous suspension is prepared by addition of water todispersible powders and granules with a dispersing or wetting agent,suspending agent one or more preservatives, and other excipients.Suspending agents include, for example, sodium carboxymethylcellulose,methylcellulose and sodium alginate. Dispersing or wetting agentsinclude naturally-occurring phosphatides, condensation products of anallylene oxide with fatty acids, condensation products of ethylene oxidewith long chain aliphatic alcohols, condensation products of ethyleneoxide with partial esters from fatty acids and a hexitol, andcondensation products of ethylene oxide with partial esters derived fromfatty acids and hexitol anydrides. Preservatives include, for example,ethyl, and n-propyl p-hydroxybenzoate. Other excipients includesweetening agents (e.g., sucrose, saccharin), flavoring agents andcoloring agents. Those skilled in the art will recognize the manyspecific excipients and wetting agents encompassed by the generaldescription above.

For oral application, tablets are prepared by mixing the active compoundwith nontoxic pharmaceutically acceptable excipients suitable for themanufacture of tablets. These excipients can be, for example, inertdiluents, such as calcium carbonate, sodium carbonate, lactose, calciumphosphate or sodium phosphate; granulating and disintegrating agents,for example, corn starch, or alginic acid; binding agents, for example,starch, gelatin or acacia; and lubricating agents, for example magnesiumstearate, stearic acid or talc. The tablets can be uncoated or they canbe coated by known techniques to delay disintegration and absorption inthe gastrointestinal tract and thereby provide a sustained action over alonger period. For example, a time delay material such as glycerylmonostearate or glyceryl distearate can be employed. Formulations fororal use can also be presented as hard gelatin capsules wherein theactive ingredient is mixed with an inert solid diluent, for example,calcium carbonate, calcium phosphate or kaolin, or as soft gelatincapsules wherein the active ingredient is mixed with water or an oilmedium, for example, peanut oil, liquid paraffin or olive oil.Formulation for oral use can also be presented as chewable gums byembedding the active ingredient in gums so that the active ingredient isslowly released upon chewing.

Additional means of systemic administration of the active compound tothe target platelets of the subject would involve a suppository form ofthe active compound, such that a therapeutically effective amount of thecompound reaches the target sites via systemic absorption andcirculation.

For rectal administration, the compositions in the form of suppositoriescan be prepared by mixing the active ingredient with a suitablenon-irritating excipient which is solid at ordinary temperatures butliquid at the rectal temperature and will therefore melt in the rectumto release the compound. Such excipients include cocoa butter andpolyethylene glycols.

The active compounds can also be systemically administered to theplatelet aggregation sites through absorption by the skin usingtransdermal patches or pads. The active compounds are absorbed into thebloodstream through the skin. Plasma concentration of the activecompounds can be controlled by using patches containing differentconcentrations of active compounds.

One systemic method involves an aerosol suspension of respirableparticles comprising the active compound, which the subject inhales. Theactive compound would be absorbed into the bloodstream via the lungs,and subsequently contact the target platelets in a pharmaceuticallyeffective amount. The respirable particles can be liquid or solid, witha particle size sufficiently small to pass through the mouth and larynxupon inhalation; in general, particles ranging from about 1 to 10microns, but more preferably 1-5 microns, in size are consideredrespirable.

Another method of systemically administering the active compounds to theplatelet aggregation sites of the subject involves administering aliquid/liquid suspension in the form of eye drops or eye wash or nasaldrops of a liquid formulation, or a nasal spray of respirable particlesthat the subject inhales. Liquid pharmaceutical compositions of theactive compound for producing a nasal spray or nasal or eye drops can beprepared by combining the active compound with a suitable vehicle, suchas sterile pyrogen free water or sterile saline by techniques known tothose skilled in the art.

Intravitreal delivery can include single or multiple intravitrealinjections, or via an implantable intravitreal device that releasesP2Y₁₂ antagonists in a sustained capacity. Intravitreal delivery canalso include delivery during surgical manipulations as either an adjunctto the intraocular irrigation solution or applied directly to thevitreous during the surgical procedure.

For systemic administration, plasma concentrations of active compoundsdelivered can vary according to compounds; but are generally1×10⁻¹⁰-1×10⁻⁴ moles/liter, and preferably 1×10⁻⁸-8×10⁻⁵ moles/liter.

The pharmaceutical utility of P2Y₁₂ antagonist compounds of thisinvention is indicated by their inhibition of ADP-induced plateletaggregation. This widely used assay, as described in S. M. O. Hourani etal. Br. J. Pharmacol. 105, 453-457 (1992) relies on the measurement ofthe aggregation of a platelet suspension upon the addition of anaggregating agent such as ADP.

The invention is illustrated further by the following examples that arenot to be construed as limiting the invention in scope to the specificprocedures described in them.

EXAMPLES Example 1 Preparation of 5′-aryl ether derivatives5-Amino-2-{2-benzyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-benzoicacid (2)

Adenosine (10 g, 37 mmol was dissolved in N,N-dimethyl formamide (100mL) and dimethoxypropane (25 mL) followed by addition of Amberlyst 15H⁺resin. The mixture was stirred 3 h at 55° C. The resin was removed byfiltration and the solvents removed in vacuo, affording2′,3′-di-O-isopropylidene adenosine (11 g, 95%).

This product (6 g, 20 mmol) was dissolved in N,N-dimethyl formamide (22mL) and stirred with triisopropylsilyl chloride and imidazole 16 h at23° C. The solution was partitioned between ether (200 mL) and brine(100 mL) and the ether phase washed with additional brine (2×50 mL). Theether was dried over magnesium sulfate and evaporated, affording5′-O-triisopropylsilyl-2′,3′-di-O-isopropylidene adenosine.

This residue was dissolved in toluene (20 mL) and treated withphenylisocyanate (3.6 g, 30 mmol) for 16 h at 25° C. A solution ofsodium bicarbonate (1 mL of 10 M) was added and the mixture evaporatedto dryness. The residue was partitioned between ethyl acetate (100 mL)and water (25 mL). The organic phase was dried with magnesium sulfateand evaporated to dryness. The solid was dissolved in tetrahydrofuran(20 mL) and stirred with tetrabutyl ammonium fluoride in tetrahydrofuran(20 mL of a 1 M solution) for 1 h in a dry ice/acetone bath. Removal ofthe solvent in vacuo followed by washing with hexane afforded the5′-alcohol (5.3 g).

A portion of the above phenylurea product (0.41 g, 0.96 mmol) wassuspended in 25 mL of 20% aqueous acetic acid and 5 mL oftetrahydrofuran/dioxane (1:1) and was stirred at 50° C. for 24 h. Thewhite suspension became a clear yellow solution. The mixture wasconcentrated and then lyophilized, to give 0.360 g (97% yield) of1-[9-(3,4-dihydroxy-5-hydroxymethyl-tetrahydro-furan-2-yl)-9H-purin-6-yl]-3-phenyl-ureaas a yellow solid. MW calculated for C₁₇H₁₈N₆O₅ (MH⁺) 387, found 387 byLCMS.

A small amount of 4A flame dried molecular sieves (cooled down by a flowof argon) was added to a vial containing a portion of the productimmediately above (0.131 g, 0.34 mmol). The mixture was capped with arubber septum and cooled down to 0° C. To this mixture trifluoroaceticacid (2.5 mL) was added via syringe and the mixture stirred at thistemperature for 15 min. Phenyl acetaldehyde dimethylacetal (0.230 ml, 4eq.) was added dropwise and the mixture stirred at 0° C. for 2 h. Onemore equivalent of phenyl acetaldehyde dimethyl acetal was added andstirred an additional five hours. The volatiles were evaporated off andthe residue was purified by flash chromatography (hexane:ethyl acetate,8:2, 1% triethylamine) to give 0.095 g of product (60% yield) as ayellow solid. MW calculated for C₂₅H₂₄N₆O₅ (MH⁺) 489, found 489 by LCMS.

A portion of this acetal product (0.068 g, 0.14 mmol) was dissolved indry N,N-dimethyl formamide (2.5 mL) and potassium tert-butoxide (0.084g, 5 eq) was added to give a yellow solution. To this mixture was added2-fluoro-5-nitrobenzoic acid (0.046 g, 1.8 eq). After 2.5 h of stirringat room temperature the mixture was concentrated and purified bypreparative HPLC to give the nucleoside analog as a white powder. MWcalculated for C₃₂H₂₇N₇O₉ (MH⁺) 654, found 654 by LCMS.

The nitro group of the product immediately above was reduced under ahydrogen atmosphere with a catalytic amount of 10% Pd/C in methanolduring 6 h. Filtration through Celite followed by HPLC purificationyielded 52 mg (62% yield) of the title compound as clear semisolid.

Example 2 Preparation of 5′-heteroaryl ether derivatives2-{2,2-Dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinicacid

To a stirred solution of1-[9-(6-hydroxymethyl-2,2-dimethyl-tetrahydro-furo[3,4-d][1,3]dioxol-4-yl)-9H-purin-6-yl]-3-phenyl-urea(43 mg, 0.1 mmol) in dry N,N′-dimethylformamide (1 mL) at 23° C. wasadded potassium tert-butoxide (45 mg, 0.4 mmol). After 30 min,2-chloronicotinic acid (60 mg, 0.4 mmol) was added to the solution. Theresulting mixture was stirred at 23° C. for 15 h and then quenched withwater (1 mL), suspended in ethyl acetate (50 mL), washed with brine(3×), dried over anhydrous sodium sulfate, filtered and concentrated invacuo to give the crude product. Preparative reverse-phase HPLC was usedto obtain the pure compound (15 mg, 27% yield) as a white solid. ¹H NMR(300 MHz, DMSO-d₆) δ 11.76 (s, 1H), 10.15 (s, 1H), 9.16 (s, 1H), 8.65(s, 1H), 8.05 (dd, J=8.5 Hz, 3.5 Hz, 1H), 7.94 (dd, J=9.0 Hz, 3.0 Hz,1H), 7.60 (d, J=2.0 Hz, 1H), 7.57 (d, J=2.0 Hz, 1H), 7.32 (t, J=12.5 Hz,2H), 7.05 (t, J=12.5 Hz, 1H), 6.96 (dd, J=12.5 Hz, 8.5 Hz, 1H), 6.25 (d,J=6.0 Hz, 1H), 5.80 (t, J=7.5 Hz, 1H), 5.10 (dd, J=9.0 Hz, 2.5 Hz, 1H),4.67 (m, 1H), 4.53 (dd, J=20.0 Hz, 4.0 Hz, 1H), 4.36 (dd, J=20.0 Hz, 4.0Hz, 1H), 1.59 (s, 3H), 1.38 (s, 3H). MW calculated for C₂₆H₂₅N₇O₇ (MH⁺)548, found 548 by LCMS.

Similarly, other 5′-substituted pyridines were prepared:

6-{2,2-Dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinicacid: (9 mg, 8% yield) as a white solid. ¹H NMR (300 MHz, DMSO-d₆) δ11.68 (s, 1H), 10.20 (s, 1H), 8.71 (s, 1H), 8.64 (d, J=3.0 Hz, 2H), 8.12(dd, J=15.0 Hz, 3.5 Hz, 1H), 7.63 (d, J=2.0 Hz, 1H), 7.61 (d, J=1.5 Hz,1H), 7.36 (t, J=13.0 Hz, 2H), 7.09 (t, J=12.5 Hz, 1H), 6.84 (dd, J=13.5Hz, 1.0 Hz, 1H), 6.33 (d, J=3.5 Hz, 1H), 5.61 (dd, J=10.0 Hz, 4.0 Hz,1H), 5.20 (dd, J=10.0 Hz, 4.5 Hz, 1H), 4.64 (m, 1H), 4.54 (m, 2H), 1.60(s, 3H), 1.40 (s, 3H). MW calculated for C₂₆H₂₅N₇O₇ (MH⁺) 548, found 548by LCMS.

6-{2,2-Dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-pyridine-2-carboxylicacid: (5 mg, 4.6% yield) as a white solid. ¹H NMR (300 MHz, DMSO-d₆) δ11.64 (s, 1H), 10.15 (s, 1H), 8.68 (s, 1H), 8.60 (s, 1H), 7.83 (dd,J=13.5 Hz, 12.5 Hz, 1H), 7.62 (m, 3H), 7.34 (t, J=12.5 Hz, 2H), 7.06 (t,J=12.5 Hz, 1H), 6.96 (dd, J=13.0 Hz, 1.0 Hz, 1H), 6.30 (d, J=4.0 Hz,1H), 5.59 (dd, J=10.0 Hz, 4.5 Hz, 1H), 5.17 (dd, J=10.0 Hz, 4.5 Hz, 1H),4.64 (m, 1H), 4.50 (m, 2H), 1.58 (s, 3H), 1.38 (s, 3H). MW calculatedfor C₂₆H₂₅N₇O₇ (MH⁺) 548, found 548 by LCMS.

6-Chloro-2-{2,2-dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinicacid (isomer A) &2-Chloro-6-{2,2-dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinicacid (isomer B):

Two isomer were obtained from the reaction with 2,6-dichloronicotinicacid. The major product was A (80 mg, 69% yield). LC-MS calculated forC₂₆H₂₄ClN₇O₇ (MH⁺) 582, found 582. The minor product was B (20 mg, 17%yield). MW calculated for C₂₆H₂₄ClN₇O₇ (MH⁺) 582, found 582 by LCMS.

6-{2,2-dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinicacid: (9 mg, 8% yield) as a white solid. ¹H NMR (300 MHz, DMSO-d₆) δ11.68 (s, 1H), 10.20 (s, 1H), 8.71 (s, 1H), 8.64 (d, J=3.0 Hz, 2H), 8.12(dd, J=15.0 Hz, 3.5 Hz, 1H), 7.63 (d, J=2.0 Hz, 1H), 7.61 (d, J=1.5 Hz,1H), 7.36 (t, J=13.0 Hz, 2H), 7.09 (t, J=12.5 Hz, 1H), 6.84 (dd, J=13.5Hz, 1.0 Hz, 1H), 6.33 (d, J=3.5 Hz, 1H), 5.61 (dd, J=10.0 Hz, 4.0 Hz,1H), 5.20 (dd, J=10.0 Hz, 4.5 Hz, 1H), 4.64 (m, 1H), 4.54 (m, 2H), 1.60(s, 3H), 1.40 (s, 3H). MW calculated for C₂₆H₂₅N₇O₇ (MH⁺) 548, found 548by LCMS.

6-{2,2-Dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-pyridine-2-carboxylicacid: (5 mg, 4.6% yield) as a white solid. ¹H NMR (300 MHz, DMSO-d₆) δ11.64 (s, 1H), 10.15 (s, 1H), 8.68 (s, 1H), 8.60 (s, 1H), 7.83 (dd,J=13.5 Hz, 12.5 Hz, 1H), 7.62 (m, 3H), 7.34 (t, J=12.5 Hz, 2H), 7.06 (t,J=12.5 Hz, 1H), 6.96 (dd, J=13.0 Hz, 1.0 Hz, 1H), 6.30 (d, J=4.0 Hz,1H), 5.59 (dd, J=10.0 Hz, 4.5 Hz, 1H), 5.17 (dd, J=10.0 Hz, 4.5 Hz, 1H),4.64 (m, 1H), 4.50 (m, 2H), 1.58 (s, 3H), 1.38 (s, 3H). MW calculatedfor C₂₆H₂₅N₇O₇ (MH⁺) 548, found 548 by LCMS.

6-Chloro-2-{2,2-dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-5-fluoro-nicotinicacid: (35 mg, 19% yield) as a white solid. MW calculated forC₂₆H₂₃ClFN₇O₇ (MH⁺) 600, found 600 by LCMS.

1-{9-[6-(3-Hydroxy-pyridin-2-yloxymethyl)-2,2-dimethyl-tetrahydro-furo[3,4-d][1,3]dioxol-4-yl]-9H-purin-6-yl}-3-phenyl-urea

Dry argon was bubbled through a solution of1-{9-[6-(3-Benzyloxy-pyridin-2-yloxymethyl)-2,2-dimethyl-tetrahydro-furo[3,4-d][1,3]dioxol-4-yl]-9H-purin-6-yl}-3-phenyl-urea(18 mg, 0.03 mmol, prepared as above) in methanol/ethyl acetate (1:1v/v, 10 mL) at 23° C. for 10 min. Palladium on activated carbon (10%)was added and the suspension was degassed by argon for another 5 min.Hydrogen (H₂) was conducted to the solution via a balloon, and thereaction proceeded for 5 h. The mixture was filtered and concentrated invacuo to give the crude product. Preparative reverse-phase HPLC was usedto obtain the pure compound (5 mg, 31% yield) as a white solid. MWcalculated for C₂₅H₂₅N₇O₆ (MH⁺) 520, found 520 by LCMS.

2-{2-Benzyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinicacid

2-{2,2-Dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinicacid (180 mg, 0.33 mmol) was dissolved in a mixture of trifluoroaceticacid and water (TFA/H₂O, 4:1 v/v, 20 mL), and the suspension was stirredat 23° C. for 30 min. The solvents were removed under reduced pressureto give the intermediate diol product (120 mg, 72% yield) as a whitesolid. MW calculated for C₂₃H₂₁N₇O₇ (MH⁺) 508, found 508 by LCMS.

To a stirred solution of the diol immediately above (0.23 mmol) in drytrifluoroacetic acid (5 mL) at 23° C. was added phenyl acetaldehyde (130mg, 1.1 mmol). The resulting mixture was stirred at 23° C. for 6 h.After the removal of most trifluoroacetic acid by evaporation underreduced pressure, the reaction was quenched with saturated sodiumbicarbonate solution (10 mL). The product was extracted using ethylacetate (50 mL), washed with brine (3×), dried over anhydrous sodiumsulfate, filtered and concentrated in vacuo to give the crude product.Preparative reverse-phase HPLC was used to obtain the pure acetalcompound (7 mg, 5% yield) as a white solid. ¹H NMR (300 MHz, DMSO-d₆) δ11.67 (s, 1H), 10.50 (s, 1H), 8.68 (s, 1H), 8.51 (s, 1H), 8.27 (dd,J=8.0 Hz, 3.0 Hz, 1H), 8.10 (dd, J=10.0 Hz, 4.0 Hz, 1H), 7.60 (dd,J=14.0 Hz, 2.0 Hz, 1H), 7.30 (m, 7H), 7.08 (m, 3H), 6.21 (d, J=4.5 Hz,1H), 5.51 (dd, J=10.5 Hz, 4.0 Hz, 1H), 5.32 (t, J=8.0 Hz, 1H), 5.06 (dd,J=10.0 Hz, 3.0 Hz, 1H), 4.70 (m, 1H), 4.62 (dd, J=20.0 Hz, 5.0 Hz, 1H),4.45 (dd, J=20.0 Hz, 5.0 Hz, 1H), 3.10 (d, J=8.5 Hz, 2H). MW calculatedfor C₃₁H₂₇N₇O₇ (MH⁺) 610, found 610 by LCMS.

Other 5′-substituted pyridine-ethers were transformed to various2′,3′-acetals using methods similar to those given immediately above.

Example 3 Synthesis of 5′-isoxazole derivatives3-[6-(6-Chloro-purin-9-yl)-2,2-dimethyl-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy]-isoxazole-5-carboxylicacid methyl ester

To a solution of[6-(6-Chloro-purin-9-yl)-2,2-dimethyl-tetrahydro-furo[3,4-d][1,3]dioxol-4-yl]-methanol(0.570 g, 1.74 mmol) in 16 mL of dry dichloromethane was addedpolymer-bound triphenylphosphine (PS-TPP; Argonaut Tech., 2.14 mmol/g,0.91 g, 1.2 eq), followed by methyl-3-hydroxy-5-isoxazolecarboxylate(0.248 g, 1 eq). The mixture was sonicated for 15 minutes then stirredat room temperature for 1 h under argon. The reaction mixture was cooledto 0° C. and under argon flow diethylazodicarboxylate (0.33 g, 1.1 eq),dissolved in 1 ml of dichloromethane, was added dropwise via syringe.The mixture was protected from light and stirred at 0° C. for 30 min.then at room temperature for 20 h. The resin was washed liberally withdichloromethane and methanol. The organic solution obtained from thewashes was concentrated to give, after flash chromatographypurification, 0.740 g of the product as a white solid (95% yield). ¹HNMR (300 MHz, CDCl₃) δ 8.76 (s, 1H), 8.23 (s, 1H), 6.42 (s, 1H), 6.23(d, J=2.1 Hz, 1H), 5.43 (dd, J=2.1 Hz, 1H), 5.14 (dd, J=3.6 Hz, 1H), 4.7(m, 1H), 4.61 (dd, J=3.9 Hz, 1H), 4.49 (dd, J=5.7 Hz, 1H), 3.93 (s, 3H),1.65 (s, 3H), 1.42 (s, 3H). MW calculated for C₁₈H₁₈ClN₅O₇ (MH⁺) 452,found 452 by LCMS.

3-{6-[6-(N-Benzyl-N-methyl-amino)-purin-9-yl]-2,2-dimethyl-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-isoxazole-5-carboxylicacid

To a solution of3-[6-(6-Chloro-purin-9-yl)-2,2-dimethyl-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy]-isoxazole-5-carboxylicacid methyl ester (0.106 g, 0.23 mmol) in tetrahydrofuran (1.2 mL) wasadded polymer-bound diisopropylethylamine (PS-DIEA; Argonaut Tech., 3.83mmol/g, 0.190 g, 3 eq), followed by addition of 0.040 mL ofN-methyl-N-benzylamine (1.2 eq). The resulting mixture was stirred atroom temperature overnight. The PS-DIEA resin was washed three timeswith dichloromethane and the solution obtained from the washes wasconcentrated to yield3-{6-[6-(N-Benzyl-N-methyl-amino)-purin-9-yl]-2,2-dimethyl-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-isoxazole-5-carboxylicacid methyl ester as yellow oil. This material (0.23 mmol) was dissolvedin 1,4-dioxane (1.2 mL) and 0.250 mL of an aqueous 2M lithium hydroxidesolution added. The mixture was stirred at room temperature for 4 h. Thecrude product, isolated after acid workup, was used without furtherpurification in the following step. MW calculated for C₂₅H₂₆N₆O₇ (MH⁺):523, found 523 by LCMS.

3-{2-Benzyl-6-[6-(N-methyl-N-benzyl-amino)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-isoxazole-5-carboxylicacid

A solution of the above acetonide (0.075 g, 0.140 mmol) dissolved in 1.5mL of dry dichloromethane was cooled in an ice bath to 0° C. To thisclear mixture was added 1.8 mL of trifluoroaceic acid. The mixture wasstirred at 0° C. for 5 h to afford the intermediate diol as a yellowsemisolid after evaporative workup. This crude product (0.23 mmol) wasdissolved at 0° C. in 1 mL of dry trifluoroacetic acid in the flask. Tothis mixture was added a small amount of 3A molecular sieves (previouslydried by flame in situ and cooled by a stream of argon). The flask wascapped with a rubber septum and cooled to 0° C., then 0.1 mL of phenylacetaldehyde dimethyl acetal was added and the mixture stirred for 18 hat 0° C. At that point, 0.1 mL more of acetal was added and stirred anadditional 6 h. Purification by HPLC yielded 52 mg of the desiredproduct as a clear semisolid (62% yield). MW calculated for C₃₀H₂₈N₆O₇(MH⁺): 585, found 585 by LCMS.

Example 4 Solution phase synthesis of 5′-carboxamide adenosine analogs2′,3′-O-Isopropylideneadenosine-5′-carboxylic acid

In a reaction vessel, bis-acetoxyiodobenzene (BAIB, 1.15 g, 3.58 mmol),2,2,6,6-tetramethyl-1-piperidinyloxy (TEMPO, 0.051 g, 0.325 mmol), and2′,3′-isopropylideneadenosine (0.500 g, 1.63 mmol) were combined, and 3mL of 1:1 acetonitrile: water mixture was added to the reaction vessel.The reaction mixture was stirred at ambient temperature under argon for1 h then filtered. The white crystalline product was washed withacetonitrile:water mixture (1:1) and dried in vacuo, to yield 0.464 g ofproduct, 89%. ¹H NMR (300 MHz, CD₃SOCD₃) δ 12.77 (br s, 1 H), 8.22 (s, 1H), 8.06 (s, 1 H), 7.27 (s, 2 H), 6.32 (s, 1 H), 5.52 (dd, J1=5.7 Hz,J2=1.8 Hz, 1 H), 5.45 (d, J=9.5 Hz, 1 H), 4.67 (d, J=1.5 Hz, 1 H), 1.52(s, 3 H), 1.35 (s, 3 H).

2-({2,2-Dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxole-4-carbonyl}-amino)-benzoicacid methyl ester

To a vial containing 2′,3′-O-Isopropylideneadenosine-5′-carboxylic acid(0.241 g, 0.75 mmol) was added 3-amino-benzoic acid methyl ester (0.144g, 0.75 mmol) in one portion at RT followed by heating at 50° C.overnight. The reaction mixture was diluted with 100 ml of ethylacetate, washed with 1N hydrochloric acid, saturated sodium bicarbonate,brine, and dried over MgSO₄. Upon removal of solvent, the solid residuewas purified with chromatography with 2-5% methanol in dichloromethaneto give 90 mg (26%) of the carboxamide as white solid. MW calculated forC₂₁H₂₂N₆O₆ (MH⁺) 455, found 455 by LCMS. This carboxamide product (90mg, 0.20 mmol) was dissolved in DMF (2 mL) and added to a flaskcontaining phenyl isocyanate (35 mg, 0.30 mmol) in 2 ml of toluene at50° C. The reaction was stirred at 50° C. overnight. Additional phenylisocyanate (35 mg, 0.20 mmol) was then added in several portions untilnearly complete consumption of the starting material was noted by TLCanalysis. The reaction mixture was then diluted with ethyl acetate (100ml), washed with saturated sodium bicarbonate, brine, and dried overmagnesium sulfate. The crude product was purified by a chromatographywith 0-2% methanol in dichloromethane to give 57 mg (50%) of pureproduct and recovered starting material (10 mg). ¹H NMR (300 MHz, CDCl₃)δ 11.53 (s, 1H), 9.21 (s, 1H), 8.54 (s, 1H), 8.36 (s, 1H), 8.21 (s, 1H),7.72 (m, 1H), 7.58 (m, 2H), 7.38 (m, 2H), 7.24 (m, 1H), 7.12 (m, 1H),6.29 (d, J=2.1 Hz, 1H), 5.66 (dd, J=6.3, 1.5 Hz, 1H), 5.56 (dd, J=6.3,1.5 Hz, 1H), 4.88 (d, J=1.8 Hz, 1H), 3.78 (s, 3H), 1.65 (s, 3H), 1.44(s, 3H). MW calculated for C₂₈H₂₇N₇O₇ (MH⁺): 574, found 574 by LCMS.

3-({2,2-Dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxole-4-carbonyl}-amino)-benzoicacid

To a vial containing 2′,3′-O-Isopropylideneadenosine-5′-carboxylic acid(250 mg, 0.778 mmol), triethylamine (1.57 mg, 1.56 mmol), and3-amino-benzoic acid allyl ester (0.276 mg, 1.56 mmol) in 0.5 ml ofN,N-dimethyl formamide at 0° C. was added PyBOP (0.443 mg, 0.856 mmol)in one portion. Reaction was continued at 0° C. for 4 h and at ambientfor 4 h. Additional PyBOP (50 mg) was added and the reaction continuedovernight at ambient temperature. The reaction mixture was diluted withethyl acetate (100 ml), washed with saturated sodium bicarbonate, brine,and dried over magnesium sulfate. The crude was purified by achromatography with 0-2% methanol in dichloromethane to give the desiredamide product. MW calculated for C₂₃H₂₄N₆O₆ (MH⁺) 481, found 481 byLCMS.

Phenyl isocyanate was coupled with the amide product using a methodsimilar to those described above, affording the intermediate phenylureacompound. MW calculated for C₃₀H₂₉N₇O₇ (MH⁺) 600, found 600 by LCMS.

This phenylurea compound (36 mg, 0.057 mmol) and morpholine (0.015 mg,0.17 mmol) were dissolved in tetrahydrofuran (5 ml) followed by additionof tetrakis(triphenylphosphine)palladium (5 mg, 0.004 mmol). Reactionwas completed in 4 h at RT. After removal of the solvent, the crudeproduct mixture was purified by preparative HPLC to give the desiredproduct. ¹H NMR (300 MHz, CDCl₃) δ 11.52 (s, 1H), 9.99 (s, 1H), 9.60 (s,1H), 8.61 (s, 1H), 8.42 (s, 1H), 7.94 (s, 1H), 7.72 (m, 4H), 7.34 (m,2H), 7.23 (m, 1H), 7.06 (m, 1H), 6.56 (s, 1H), 5.56 (m, 2H), 4.88 (s,1H), 1.58 (s, 3H), 1.39 (s, 3H).

Example 5 Solution phase synthesis of a sulfonamideN-{2,2-Dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxole-4-carbonyl}-methanesulfonamide

To a vial containing 2′,3′-O-Isopropylideneadenosine-5′-carboxylic acid(20 mg, 0.045 mmol), dimethylaminopyridine (5 mg, 0.045 mmol), andmethanesulfonamide (0.009 mg, 0.091 mmol) in dichloromethane (0.5 ml)was added dicyclohexylcarbodiimide (10 mg, 0.05 mmol). The mixture wasstirred for 2 days at RT. Additional dicyclohexylcarbodiimide (10 mg,0.05 mmol) and dimethylaminopyridine (5 mg, 0.045 mmol) were added andthe reaction was continued at RT overnight. To the reaction mixture wasadded ethyl acetate (75 ml), which was washed with 1 N hydrochloricacid, water, saturated sodium bicarbonate, brine, and dried overmagnesium sulfate. Upon removal of solvent, the residue was purified byprep. HPLC to yield 13 mg of desired product (55%). MW calculated forC₂₁H₂₃N₇O₇S (MH⁺) 518, found 518 by LCMS.

Example 6 Solid phase synthesis of ureas and acetals from polymer bound5′-proline-amides of adenosine

Commercially available hydroxymethylsulfanylmethyl (HESM) polystyreneresin (1.4 mmol/g, 200 mesh, NovaBiochem; 2.82 g, 3.95 mmol) was swelledfor 15 minutes in 50 mL. In a separate reaction vessel, Boc-Pro-OH (3.40g, 15.8 mmol), HATU (5.7 g, 15.0 mmol), dimethylaminopyridine (0.24 g,1.98 mmol), and diisopropylethylamine (3.5 mL, 19.8 mmol) were dissolvedin 40 mL of N,N-dimethyl formamide and stirred for 15 minutes.N,N-dimethyl formamide was drained from the HESM resin and the solutionof activated proline derivative was added to the resin. The resin wasagitated at RT for 17 h. The solvent was then drained and the resinwashed with N,N-dimethyl formamide (3×30 mL), dichloromethane (3×30 mL),methanol (3×30 mL), dichloromethane (2×30 mL), methanol (3×30 mL) anddried in vacuo overnight. Mass of resin: 3.42 g, 93% loading.

Removal of BOC Protecting Group:

Resin obtained in the previous step was agitated with a 40%trifluoroacetic acid/1.0 dichloromethane solution (75 mL) for 15minutes. The solvent was drained, and a fresh solution of 40%trifluoroacetic acid in dichloromethane was added, and resin wasagitated for another 15 minutes. After this the resin was washed withdichloromethane (5×40 mL), 20% diisopropylethylamine/dichloromethane(2×30 mL), dichloromethane (3×30 mL), and methanol (5×40 mL). The resinwas dried under vacuum. A chloranil test indicated the presence of afree amino group, and this proline-bound resin was carried over to thenext step.

The proline resin product from the previous step was swelled in 50 mLN,N-dimethyl formamide for 30 minutes, after that the N,N-dimethylformamide was drained. In a separate reaction vessel,2′,3′-O-Isopropylideneadenosine-5′-carboxylic acid (1.40 g, 4.35 mmol),dichloroethane (0.91 g, 4.74 mmol), HOBtH₂O (0.73 g, 4.74 mmol), anddiisopropylethylamine (3.5 mL, 19.8 mmol) were dissolved in 55 mL ofN,N-dimethyl formamide. The solution was stirred for 15 minutes, andthen added to the proline resin. The resin was agitated at RT for 17 h.The solvent was drained, the resin was washed with N,N-dimethylformamide (3×30 mL), dichloromethane (3×30 mL), methanol (3×30 mL),dichloromethane (2×30 mL), methanol (3×30 mL), and dried in vacuo for 48h. A chloranil test performed on a few sample beads indicated thatcoupling had occurred. A small amount of resin was cleaved using thefollowing procedure to verify attachment of the carboxylic acid, andanalysed by LCMS. Calculated mass for C₁₈H₂₂N₆O₆ (MH⁺): 419, found 419by LCMS. Mass of resin: 3.66 g, 0.55 mmol/g, 82% in three steps.

General Cleavage Procedure for Analysis of HESM Resin:

A small amount of resin is suspended in a solution of 5-6 equivalents ofm-chloroperbenzoic acid in dichloromethane and agitated for 7-8 hours atRT. The solution is then drained, and the resin is washed 5-6 times withfresh dichloromethane. Then resin is suspended in a solution of 4-5equivalents of DBU in dichloromethane, and agitated at RT for 4-5 hours.The resin is then filtered and the solution is analyzed by LC/MS, HPLCor another method. The compounds are recovered from solution by rotaryevaporation.

The adenosine-proline amide-derivatized resin from the previousresin-synthesis step (0.5 g, 0.275 mmol) was suspended in anhydrousN,N-dimethyl formamide (10 mL). Ethyl isocyanate was added (0.43 mL, 5.5mmol), and the reaction mixture was heated in a capped vial at 55° C.for 16 h. The resin was then drained and washed with N,N-dimethylformamide (3×10 mL), dichloromethane (3×10 mL), methanol (3×10 mL),dichloromethane (2×10 mL), methanol (3×10 mL), and the procedure wasrepeated once again then the resin dried in vacuo for 24 h. A negativechloranil test indicated complete reaction. A small amount of this resin(bound to the 5′-adenosine-(2′,3′-acetonide)-proline amide-derivatizedas the 6-ethylurea) was cleaved by the procedure described for cleavageof HESM resin, and the isolated product was analyzed by LCMS. Masscalculated for C₂₁H₂₇N₇O₇ (MH⁺): 490, found 490 by LCMS. Loading 0.52mmovg, mass 0.52 g, 98%.

The procedure described above was also used to prepare N⁶-ureas withR_(d)=—C₆H₁₁, —Ph, —CH₂Ph, —CH₂CH₂Ph, -cyclopentyl, andtrans-2-phenyl-cycloprop-1-yl groups using the appropriate isocyanates.

1-{6-[6-(3-Ethyl-ureido)-purin-9-yl]-2-phenyl-tetrahydro-furo[3,4-d][1,3]dioxole-4-carbonyl}-pyrrolidine-2-carboxylicacid

The resin described above (0.05 g, 0.026 mmol) was suspended intrifluoroacetic acid, then benzaldehyde (0.095 g, 0.9 mmol) was addedall at once. The resin was agitated in a tightly closed vial for 24 h.The resin was drained, washed with dichloromethane (5×3 mL), 20%diisopropylethylamine/dichloromethane (2×3 mL), dichloromethane (3×3mL), and methanol (5×3 mL), then dried in vacuo for 3 h. The resin wascleaved using the cleavage procedure described above, the crude productwas collected, analyzed by LCMS and purified by preparative HPLC.Calculated MW for C₂₅H₂₇N₇O₇ (MH⁺): 538, found: 538 by LCMS.

The following analogs were prepared in a similar manner to thatdescribed above, using appropriate combinations of isocyanates andaldehydes. Compounds analyzed by LCMS.

-   R_(d)=ethyl, R_(a)=benzyl Calculated MW for C26H29N7O7: 552.55    (MH+), found: 552.4.-   R_(d)=ethyl, R_(a)=4-biphenyl Calculated MW for C31H31N7O7: 614.62    (MH+), found: 614.3-   R_(d)=ethyl, R_(a)=3-biphenyl Calculated MW for C31H31N7O7: 614.62    (MH+), found: 614.3-   R_(d)=ethyl, R_(a)=2-naphthyl Calculated MW for C29H29N7O7: 588.58    (MH+), found: 588.1.-   R_(d)=n-hexyl, R_(a)=phenyl Calculated MW for C29H35N7O7: 594.63    (MH+), found: 594.3.-   R_(d)=n-hexyl, R_(a)=benzyl Calculated MW for C30H37N7O7: 608.65    (MH+), found: 608.2.-   R_(d)=n-hexyl, R_(a)=4-biphenyl Calculated MW for C35H39N7O7: 670.73    (MH+), found: 670.3.-   R_(d)=n-hexyl, R_(a)=3-biphenyl Calculated MW for C35H39N7O7: 670.73    (MH+), found: 670.3.-   R_(d)=n-hexyl, R_(a)=2-naphthyl Calculated MW for C33H37N7O7: 644.69    (MH+), found: 644.3.-   R_(d)=cyclopentyl, R_(a)=benzyl Calculated MW for C29H33N7O7: 592.62    (MH+), found: 592.3.-   R_(d)=cyclopentyl, R_(a)=phenyl Calculated MW for C28H31N7O7: 578.59    (MH+), found: 578.3.-   R_(d)=cyclopentyl, R_(a)=4-biphenyl Calculated MW for C34H35N7O7:    654.68 (MH+), found: 654.3.-   R_(d)=cyclopentyl, R_(a)=3-biphenyl Calculated MW for C34H35N7O7:    654.68 (MH+), found: 654.3.-   R_(d)=cyclopentyl, R_(a)=2-naphthyl Calculated MW for C32H33N7O7:    628.65 (MH+), found: 628.4.-   R_(d)=benzyl, R_(a)=benzyl Calculated MW for C31H31N7O7: 614.62    (MH+), found: 614.3.-   R_(d)=benzyl, R_(a)=phenyl Calculated MW for C30H29N7O7: 600.59    (MH+), found: 600.3.-   R_(d)=benzyl, R_(a)=2-naphthyl Calculated MW for C34H31N7O7: 650.65    (MH+), found: 650.3.-   R_(d)=benzyl, R_(a)=4-biphenyl Calculated MW for C36H33N7O7: 676.69    (MH+), found: 676.3.-   R_(d)=benzyl, R_(a)=3-biphenyl Calculated MW for C36H33N7O7: 676.69    (MH+), found: 676.3.-   R_(d)=ethylphenyl, R_(a)=benzyl Calculated MW for C32H33N7O7: 628.65    (MH+), found: 628.4.-   R_(d)=ethylphenyl, R_(a)=phenyl Calculated MW for C31H31N7O7: 614.62    (MH+), found: 614.5.-   R_(d)=ethylphenyl, R_(a)=4-biphenyl Calculated MW for C37H35N7O7:    690.72 (MH+), found: 690.4.-   R_(d)=ethylphenyl, R_(a)=3-biphenyl Calculated MW for C37H35N7O7:    690.72 (MH+), found: 690.5.-   R_(d)=ethylphenyl, R_(a)=2-naphthyl Calculated MW for C35H33N7O7:    664.68 (MH+), found: 664.4.-   R_(d)=cyclopropyl-trans-2-phenyl, R_(a)=benzyl Calculated MW for    C33H33N7O7: 640.66 (MH+), found: 640.3.-   R_(d)=cyclopropyl-trans-2-phenyl, R_(a)=3-biphenyl Calculated MW for    C38H35N7O7: 702.73 (MH+), found: 702.6.-   R_(d)=cyclopropyl-trans-2-phenyl, R_(a)=2-naphthyl Calculated MW for    C36H33N7O7: 676.69 (MH+), found: 676.6.-   R_(d)=phenyl, R_(a)=benzyl Calculated MW for C30H29N7O7: 600.59    (MH+), found: 600.3.-   R_(d)=phenyl, R_(a)=phenyl Calculated MW for C29H27N7O7: 586.57    (MH+), found: 586.2.-   R_(d)=phenyl, R_(a)=2-naphthyl Calculated MW for C33H29N7O7: 636.63    (MH+), found: 636.5.-   R_(d)=phenyl, R_(a)=4-biphenyl Calculated MW for C35H31N7O7: 662.66    (MH+), found: 662.5.-   R_(d)=phenyl, R_(a)=3-biphenyl Calculated MW for C35H31N7O7: 662.66    (MH+), found: 662.5.-   R_(d)=phenyl, R_(a)=3-thianaphthene Calculated MW for C31H27N7O7S:    642.65 (MH+), found: 642.0.-   R_(d)=ethylphenyl, R_(a)=3-thianaphthene Calculated MW for    C33H31N7O7S: 670.71 (MH+), found: 670.0.-   R_(d)=ethylphenyl, R_(a)=3-thianaphthene Calculated MW for    C32H29N7O7S: 656.68 (MH+), found: 656.1.-   R_(d)=n-hexyl, R_(a)=3-thianaphthene Calculated MW for C31H35N7O7S:    650.72 (MH+), found: 650.2.-   R_(d)=n-hexyl, R_(a)=CHCHPh Calculated MW for C31H37N7O7: 620.67    (MH+), found: 620.4.-   R_(d)=n-hexyl, R_(a)=CCPh Calculated MW for C31H35N7O7: 618.65    (MH+), found: 618.0.-   R_(d)=cyclopropyl-trans-2-phenyl, R_(a)=CCPh Calculated MW for    C34H31N7O7: 650.65 (MH+), found: 650.1.-   R_(d)=ethylphenyl, R_(a)=CCPh Calculated MW for C33H31N7O7: 638.64    (MH+), found: 638.1.-   R_(d)=benzyl, R_(a)=CCPh Calculated MW for C32H29N7O7: 624.62 (MH+),    found: 624.1.-   R_(d)=ethyl, R_(a)=CCPh Calculated MW for C27H27N7O7: 562.55 (MH+),    found: 562.0.-   R_(d)=cyclopropyl-trans-2-phenyl, R_(a)=4-biphenyl Calculated MW for    C38H35N7O7: 702.73 (MH+), found: 702.1.-   R_(d)=cyclopropyl-trans-2-phenyl, R_(a)=phenyl Calculated MW for    C32H31N7O7: 626.63 (MH+), found: 626.0.-   R_(d)=cyclopropyl-trans-2-phenyl, R_(a)=CHCHPh Calculated MW for    C34H33N7O7: 652.67 (MH+), found: 652.1.-   R_(d)=ethylphenyl, R_(a)=CHCHPh Calculated MW for C33H33N7O7: 640.66    (MH+), found: 640.3.

Example 7 Synthesis of a 5′ sulfonylurea as in Scheme 101-[9-(2-Benzyl-6-ureidomethyl-tetrahydro-furo[3,4-d][1,3]dioxol-4-yl)-9H-purin-6-yl]-3-phenyl-ureamethylsulfonamide

In a reaction vessel were combined2′,3′-O-benzylidene-N-6-(phenylurea)adenosine (0.200 g, 0.409 mmol),p-toluene sulfonyl chloride (0.148 g, 0.778 mmol), and 4.0 mL ofpyridine. The mixture was stirred for 4 h at RT. The solvent was removedin vacuo and the tosylate (0.260 g, 99%) was recovered as a yellowsolid. MW calculated for C₃₂H₃₀N₆O₇S: 643.19 (MH⁺), found 642.9.

This tosylate product (0.260 g, 0.409 mmol) was dissolved in 3 mL ofanhydrous N,N-dimethyl formamide, sodium azide (0.266 g, 4.09 mmol) wasadded, and the mixture was heated at 80° C. in a closed vial for 7 hwhile being stirred. The mixture was diluted with 50 mL ofdichloromethane and extracted with 5% sodium bicarbonate solution andbrine. The organic layer was separated, dried over anhydrous sodiumsulfate and solvent was removed in vacuo. The azide derivative wasrecovered (0.183 g, 87%) as a white solid. MW calculated for C₂₅H₂₃N₉O₄:514.19 (MH⁺), found 514.1.

This residue containing the azide (0.180 g, 0.351 mmol) was dissolved in6 mL of tetrahydrofuran/water mixture (18:1), polystyrene-boundtriphenylphospine was added (2.19 mmol/g, 0.800 g, 1.75 mmol), and thereaction mixture was stirred at RT under argon for 24 h. The reactionmixture was then filtered, the solvent was removed in vacuo and thecrude product was chomatographed on a silica gel column (2 cm×15 cm).The column was eluted with dichloromethane/methanol/triethylamine(88:10:2) to give 0.072 g (42%) of the amine as a white solid. MWcalculated for C₂₅H₂₅N₇O₄: 488.20 (MH⁺), found 488.0.

A portion of the amine product above (0.022 g, 0.046 mmol) was dissolvedin 2 mL of dichloromethane (anhydrous) and methylsulfonylethylcarbamate(0.008 g, 0.046 mmol) was added. The reaction was stirred at RT underargon for 72 h. Solvent was then removed in vacuo, and the crude productwas purified by preparative HPLC (acetonitrile/0.1% trifluoroaceticacid//water/0.1% trifluoroacetic acid buffer). 0.016 g (59%) of productwas recovered as a white solid. MW calculated for C₂₇H₂₈N₈O₇S: 609.18(MH⁺), found 609.4. ¹H NMR (300 MHz, CD₃SOCD₃) δ 11.69 (br s, 1 H),10.22 (br s, 1 H), 8.69 (s, 1 H), 8.65 (s, 1 H), 7.60 (d, J=7.8 Hz, 2H), 7.36-7.21 (m, 6 H), 7.07 (t, J=7.5 Hz, 1H), 6.90-6.87 (m, 1 H), 6.57(s, 1 H), 6.22 (d, J=2.4 Hz, 1 H), 5.75 (s, 1 H), 5.45 (dd, J1=6.3 Hz,J2=2.1 Hz, 1 H), 5.31 (t, J=5.1 Hz, 1 H), 4.92 (dd, J1=6.6 Hz, J2=3.3Hz, 1 H), 4.16-4.02 (m, 5 H), 3.11 (s, 3 H).

Example 8 5′-Homologated Derivatives as in Scheme 113-{2,2-Dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-yl}-acrylicacid methyl ester

To a vial containing the starting compound,1-[9-(6-Hydroxymethyl-2,2-dimethyl-tetrahydro-furo[3,4-d][1,3]dioxol-4-yl)-9H-purin-6-yl]-3-phenyl-urea(1.07 g, 2.5 mmol), in 10 ml of dimethylsulfoxide was added IBX (1.06 g,3.75 mmol) in one portion at RT. The white solid gradually dissolved asthe reaction proceeded. After stirring at RT for 2 hours,methyl(triphenylphosphorylidene) acetate (0.84 g, 2.5 mmol) was added inone portion. The reaction was run at RT overnight. To the reactionmixture was added ethyl acetate (100 ml), which was washed withsaturated NaHCO₃, brine, and dried over MgSO₄. After removal of solvent,the residue was recrystallized with isopropyl alcohol to provide thetitle compound. MW calculated for C₂₃H₂₄N₆O₆ (MH⁺) 481.5 found 481.3 byLCMS.

3-{2,2-Dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-yl}-propionicacid

To a round bottom flask containing product from the previous step (1.2g, 2.5 mmol) and palladium on carbon (10% w/w, 10 mg) under nitrogen wasadded methanol (20 ml). After flushing with hydrogen gas, the reactionmixture was stirred under a hydrogen atmosphere using a hydrogen balloonovernight at RT. Upon filtration and removal of solvent, the crude whitesolid product was recrystallized from isopropyl alcohol to give thedesired methyl ester compound. MW calculated for C₂₃H₂₆N₆O₆ (MH⁺) 483.5found 483.2.

The methyl ester (500 mg, 1.04 mmol) was dissolved in 4 ml of methanoland sodium hydroxide (83 mg, 2.1 mmol) was then added. The reaction wasstirred overnight at RT. After removal of methanol, acetic acid (2.1mmol, 120 mmol) was added and a white solid precipitated. The solventwas removed under vacuum. The residue was recrystallized from water toprovide the pure desired product as a white solid (0.4 g, 83%). MWcalculated for C₂₂H₂₄N₆O₆ (M−1) 467.5 found 467.4 by LCMS.

1-(3-{2,2-Dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-yl}-propion-1-yl)-pyrrolidine-2-carboxylicacid

To a vial containing the carboxylic acid product from the previous step(0.117 g, 0.25 mmol), thionyl chloride (0.3 g, 2.5 mmol) was added at 0°C. After addition, the cold bath was removed followed by addition of twodrops of N,N-dimethyl formamide. The reaction mixture was heated to 50°C. for 30 minutes. The excess of thionyl chloride was removed undervacuum and the solid residue was washed with ethyl ether to give theacid chloride. The acid chloride (61 mg, 0.125 mmol) was added to a vialcontaining L-proline methyl ester (23 mg, 0.138 mmol) and triethylamine(28 mg, 0.275 mmol) in 1 mL of dichloromethane at 0° C. The reaction wasgradually warmed to RT overnight. To the reaction mixture was addedethyl acetate (75 ml), which was washed with 1 N hydrochloric acid,saturated sodium bicarbonate, and dried over magnesium sulfate. Theresidue was purified by elution from a silica column using 2% methanolin dichloromethane to give purified prolylmethyl ester product (10 mg,14%). MW calculated for C₂₈H₃₃N₇O₇ (MH⁺) 580.6 found 580.3 by LCMS. Theprolylmethyl ester (6 mg, 0.010 mmol) was dissolved in tetrahydrofuran(0.1 ml) followed by addition of 6 μl 15% sodium hydroxide. Afterstirring at RT for 2 hrs, ethyl acetate (50 ml) was added and enough 1 Nhydrochloric acid was added to adjust the pH to 3. The organic layer waswashed with brine and dried over magnesium sulfate. After removal ofsolvent, the title compound was obtained as a white powder. MWcalculated for C₂₇H₃₁N₇O₇ (MH⁺) 566.6 found 566.3 by LCMS.

Example 9 Enzymatic synthesis of a mixture of1-Ethyl-3-[9-(6-hydroxymethyl-2-styryl-tetrahydro-furo[3,4-d][1,3]dioxol-4-yl)-9H-purin-6-yl]-ureaisomers from the corresponding isomeric mixture of 5′-AMP acetal/ureaderivatives

The 2′,3′-(cinnamylacetal)-N⁶-(ethylurea) derivative of AMP (0.750 g,0.14 mmol) as a mixture of acetal diastereoisomers was dissolved inwater (25 mL, 1.4 mol) in a round bottom flask and the pH was adjustedto 8.3 with NaOH. The temperature was adjusted to 35 C, and alkalinephosphatase (0.003 g, 0.00004 mol) was added. Within 15 minutes, themixture became rather heterogenous, and methanol (20 mL, 0.5 mol) wasadded to resolubilize the nucleoside product. After 4 h the reaction wasjudged essentially complete by HPLC. The reaction was worked up byadding more MeOH (20 mL), heating to 60 C to denature the enzyme, andfiltering through a 0.22 uM filter. The methanol was evaporated invacuo, and a white, fine-particle solid precipitated from the remainingsolvent. This mixture was cooled in an ice bath and filtered. Washingthe material with water, followed by drying over P₂O₅ in a dessicatorafforded the title product as a mixture of acetal diastereomers. Dryweight 440 mg (0.10 mmol, 71% yield).

Example 103-{6-[6-(3-Ethyl-ureido)-purin-9-yl]-2-styryl-tetrahydro-furo[3,4-d][1,3]dioxol-4-yl}-acrylicacid methyl ester

1-Ethyl-3-[9-(6-hydroxymethyl-2-styryl-tetrahydro-furo[3,4-d][1,3]dioxol-4-yl)-9H-purin-6-yl]-urea(5.0 g, 11 mmol) was suspended in dry acetonitrile (50 mL) andDess-Martin periodinate (6.7 g, 16 mmol) was added. The suspension wasstirred 2 h, after which time proton NMR of an aliquot showed completeconversion to the aldehyde.(Methoxycarbonylmethylene)triphenylphosphorane (3.9 g, 12 mmol) wasadded and stirring was continued overnight. The reaction mixture wasthen diluted with ethyl acetate (300 mL), washed with saturated sodiumbicarbonate/thiosulfate solution (100 mL), dried with sodium sulfate andfiltered. The filtrate was evaporated and the solid was dissolved in hotisopropyl alcohol (50 mL). It was allowed to cool, then heptane wasadded and it was stirred overnight. The resulting precipitate was washedwith heptane and dried under vacuum, affording the desired product (2.9g, 71%). ¹H-NMR (300 MHz, d₆ DMSO) δ 1.15 (t, 3H, J=7 Hz), 3.21 (q, 2H,J=7 Hz), 3.59 (s, 3H), 4.98 (m, 1H), 5.28 (ψt, 1H, J=6 Hz), 5.51 (dd,1H, J=6 Hz, <2 Hz), 5.70 (d, 1H, J=16 Hz), 5.90 (d, 1H, J=6 Hz), 6.30(dd, 1H, J=6 Hz, 16 Hz), 6.45 (d, 1H, J<2 Hz), 6.95 (d, 1H, J=15 Hz),7.35 (m, 3H), 7.45 (d, 2H, J=7 Hz), 8.50 (s, 1H), 8.60 (s, 1H), 9.30 (t,1H, J=6 Hz), 9.60 (s, 1H).

Example 113-{6-[6-(3-Ethyl-ureido)-purin-9-yl]-2-styryl-tetrahydro-furo[3,4-d][1,3]dioxol-4-yl}-propionicacid methyl ester

3-{6-[6-(3-Ethyl-ureido)-purin-9-yl]-2-styryl-tetrahydro-furo[3,4-d][1,3]dioxol-4-yl}-acrylicacid methyl ester (250 mg, 0.5 mmol) was dissolved in dry methanol (3mL). Copper (II) sulfate (90 mg, 0.5 mmol) was added followed by sodiumtetrahydroborate (90 mg, 2.5 mmol) and the reaction was stirred 48 h.The reaction was diluted with water, filtered, and concentrated invacuo. The residue was dissolved in ethyl acetate and precipitated withheptane. The precipitate was dissolved in dichloromethane and waschromatographed on silica gel with dichloromethane-methanol (95:5) aseluent, affording the title compound (125 mg, 50%). ¹H-NMR (300 MHz, d₆DMSO). δ 1.15 (t, 3H, J=7 Hz), 1.90 (m, 2H), 2.19 (m, 2H), 3.21 (q, 2H,J=7 Hz), 3.55 (s, 3H), 4.20 (m, 1H), 4.98 (dd, 1H, J=4 Hz, 6 Hz), 5.45(dd, 1H, J=3 Hz, 7 Hz), 5.85 (d, 1H, J=6 Hz), 6.25 (d, 1H, J=3 Hz), 6.27(dd, 1H, J=6 Hz, 16 Hz), 6.90 (d, 1H, J 16 Hz), 7.35 (m, 3H), 7.50 (d,2H, J=7 Hz), 8.56 (s, 1H), 8.57 (s, 1H), 9.30 (t, 1H, 3=5 Hz), 9.60 (s,1H).

Example 123-{6-[6-(3-Ethyl-ureido)-purin-9-yl]-2-styryl-tetrahydro-furo[3,4-d][1,3]dioxol-4-yl}-propionicacid

3-{6-[6-(3-Ethyl-ureido)-purin-9-yl]-2-styryl-tetrahydro-furo[3,4-d][1,3]dioxol-4-yl}-propionicacid methyl ester (5.0 g, 10 mmol) was dissolved in tetrahydrofuran (300mL). Water (100 mL) was added, followed by lithium hydroxide (1.0 g, 25mmol). The solution was allowed to stir 16 h at room temperature. It wasacidified to pH 5 with acetic acid, concentrated in vacuo, thenextracted with chloroform (300 mL). The organic extract was evaporated,redissolved in ethyl acetate and precipitated with heptane to afford thefinal product (3.9 g, 80%). ¹H-NMR (300 MHz, d₆ DMSO) δ 1.14 (t 3H, J=7Hz), 1.90 (m, 2H), 2.19 (m, 2H), 3.26 (q, 2H, 3=6 Hz), 4.17 (m, 1H),4.93 (ψt, 1H, J=6 Hz), 5.43 (dd, 1H, J=3 Hz, 7 Hz), 5.84 (d 1H, J=6 Hz),6.24 (d, 1H, J=3 Hz), 6.28 (dd, 1H, J=7 Hz, 13 Hz), 6.90 (d, 1H, J 16Hz), 7.35 (m, 3H), 7.51 (d, 2H, 3=7 Hz), 8.56 (s, 1H), 8.61 (s, 1H),9.30 (t, 1H, 3=6 Hz).

Example 13 Synthesis of 5′-Ethers as in Scheme 123-Chloro-2-{6-[6-(3-ethyl-ureido)-purin-9-yl]-2-trans-styryl-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxymethyl}-benzoicacid

1-Ethyl-3-[9-(6-hydroxymethyl-2-trans-styryl-tetrahydro-furo[3,4-d][1,3]dioxol-4-yl)-9H-purin-6-yl]-urea(0.200 g, 0.44 mmol) was suspended in tetrahydrofuran (5 mL) and sodiumhydride (60% w/w in oil, 0.106 g, 2.65 mmol) added. Once the bubblingceased, added methyl-2-bromomethyl-3-chloro-benzoate (0.233 g, 0.88mmol) and stirred the reaction mixture overnight at room temperature.Mass spectral analysis indicated that the reaction was complete and thatthe product was the title compound, arising from in situ hydrolysis ofthe methyl ester. The pH was lowered to 5 with acetic acid, and themixture partitioned between ethyl acetate (40 mL) and 50% saturatedsodium chloride (50 mL). The layers were separated and the ethyl acetatelayer concentrated to dryness. The residue was reconstituted in inaqueous acetonitrile and the product purified on preparative HPLC (C₁₈column, gradient from 0.05 M ammonium acetate (pH 6.5) to acetonitrileover 20 minutes). The solvent was removed from the fraction containingthe product, giving the title compound after overnight lyophilization(0.194 g, 70%)

MW calculated for C₃₀H₂₉ClN₆O₇ (MH⁻) 620.0 found 619.6 by LCMS. ¹H-NMR(300 MHz, CDCl₃) δ 1.29 (t 3H), 3.42 (q, 2H), 3.77 (dd, 1H), 3.97 (dd,1H), 4.70 (s, 1H), 4.90 (d, 1H), 5.05 (d, 1H), 5.19 (d, 1H), 5.60 (dd,1H), 5.86 (d, 1H), 6.18 (dd, 1H), 6.28 (d, 1H), 6.83 (d, 1H), 7.36 (m,7H), 7.89 (d, 1H), 8.52 (s, 1H), 8.57 (s, 1H), 9.33 (s, 1H), 9.56 (t,1H).

Example 14 Inhibition of ADP-Induced Platelet Aggregation

Isolation of Platelets: Human blood is obtained from informed healthyadult volunteers. Blood is collected into one-sixth volume ofacid/citrate/dextrose (ACD) buffer (85 mM sodium citrate, 65 mM citricacid, and 110 mM glucose). Collected blood is placed in a water bath at37° C. for 30 minutes. Blood is then centrifuged at 275×g for 16 minutesat room temperature and the platelet-rich plasma is removed andcentrifuged at 2200×g for 13 minutes at room temperature. The plateletpellet is resuspended in 40 mL of HEPES-buffered Tyrode's solution (137mM NaCl, 2.7 mM KCl, 1 mM MgCl₂, 2 mM CaCl₂, 12 mM NaHCO₃, 0.36 mMNaH₂PO₄, 5.5 mM glucose, 5 mM HEPES pH 7.4, 0.35% bovine serum albuminor 0.35% human serum albumin) containing 10 U/mL heparin and 5 μM (finalconcentration) prostaglandin I₂ (PGI₂). The platelet suspension isincubated in a 37° C. water bath for 10 minutes and then 5 μM (finalconc.) PGI₂ is added just before centrifugation at 1900×g for 8 minutes.The resulting pellet is resuspended in 40 mL of HEPES-buffered Tyrode'ssolution containing 5 μM (final concentration) PGI₂ and then isincubated for 10 minutes in a 37° C. water bath. A small aliquot (500μL) of the platelet suspension is removed for platelet counting. Priorto centrifugation 5 μM (final concentration) PGI₂ is added to thesuspension and then the suspension is centrifuged at 1900×g for 8minutes. The pellet is resuspended at a density of 5×10⁸ cells/mL inHEPES-buffered Tyrode's solution containing 0.05 U/mL apyrase.

Aggregation Studies: ADP-induced platelet aggregation is determined bymeasuring the transmission of light through a 0.5 ml suspension ofstirred (1000 rpm) washed platelets in a lumi-aggregometer at 37° C.(Chrono-Log Corp. Havertown, Pa.). The baseline of the instrument is setusing 0.5 ml of Hepes-buffered Tyrode's solution. Prior to aggregationmeasurements, the platelet suspension is supplemented with 1 mg/mlfibrinogen. Platelet aggregation is initiated by the addition ofindicated concentrations of ADP or other agonists, and the lighttransmission is continuously recorded for at least 8 min. Wheninhibitors of platelet aggregation are tested, platelets are incubatedfor 2 min in the presence of indicated concentrations of inhibitorbefore addition of ADP or other agonists, and the response is recordedfor at least 8 min. The potency of agonists and inhibitors of plateletaggregation is calculated from both the rate of aggregation and themaximal extent of aggregation obtained for each determination by fittingthe data to a four-parameter logistic equation using the GraphPadsoftware package (GraphPad Corp. San Diego, Calif.).

When a broad range of concentrations of P2Y₁₂ antagonist is tested(usually from 1 nM to 100 μM), an IC₅₀ value is also obtained. IC₅₀values represent the concentration of antagonist necessary to inhibit by50% the aggregation elicited by a given concentration of ADP.

Example 15 Inhibition of ADP-induced Platelet Aggregation in Whole Blood

Human blood is obtained from informed healthy adult volunteers. Blood iscollected into syringes containing heparin, sodium citrate, PPACK orhirudin as anticoagulant. Blood is carefully transferred to a conicaltube and maintained at room temperature. Assays are conducted within 60min from the collection of the blood sample. ADP-induced plateletaggregation is performed using the impedance mode of an aggregometer(Chrono-Log Corp. Havertown, Pa.). Blood is gently mixed and an aliquotof 500 μL is transferred to a measurement cuvette, then, 450 μL of warmsterile saline is added to each cuvette and the sample is stirred at1000 rpm. The impedance probe is introduced into the cuvette and thesample is allowed to warm for approx. 3-4 minutes in the aggregometer.The basal impedance is recorded for 1 minute and then 50 μL of theappropriate concentrations of ADP are added to generate an ADP doseresponse curve. For the evaluation of P2Y₁₂ receptor antagonists onplatelet aggregation, after the basal impedance is recorded for 1 minuteas indicated above, blood samples are supplemented with 50 μL of theantagonist or vehicle and after 2 minutes, 50 μL of ADP (EC₉₀; usually5-10 μmol/L ADP) are added and the impedance is recorded for up to 8minutes. The potency of agonists and inhibitors of platelet aggregationis calculated from the impedance values obtained in each sample byfitting the data to a four-parameter logistic equation using theGraphPad software package (GraphPad Corp. San Diego, Calif.).

Example 16 Effects on Platelet Aggregation in vivo

To evaluate the ability of these compounds to inhibit plateletaggregation in vivo, an experimental protocol similar to the method ofR. G. Humphries et al. (Br. J. Pharmacol. 115:1110-1116, 1995) isperformed.

Surgical Preparation and Instrumentation: Male Sprague-Dawley rats areanesthetized. Body temperature is maintained at 37±0.5° C. with aheating lamp. Animals breathe spontaneously and a tracheotomy isperformed to ensure a patent airway. A cannula containing heparinizedsaline is introduced into the left femoral artery and connected to atransducer to record blood pressure and heart rate. Cannulae containingnon-heparinized saline are introduced into the left common carotidartery and left jugular vein for withdrawal of arterial blood samplesand intravenous administration of compounds, respectively.

Experimental Protocol: Either compound or vehicle is administered toeach animal as an infusion. Blood samples are taken immediately prior tothe first infusion, at the end of each infusion and 20 min aftercessation of the final infusion for measurement of platelet aggregationex vivo. Immediately after sampling, ADP-induced platelet aggregation ismeasured in duplicate in 0.5 ml blood samples diluted 1:1 with salineand incubated at 37° C. for 4 min. For the final minute of this period,cuvettes are transferred to a lumi-aggregometer and the sample stirredat 900 rpm. ADP (3 μM) is added in a volume of 20 μl and the aggregationresponse is recorded using the impedance mode of the aggregometer.

Example 17 Inhibition of Thrombus Formation in Anesthetized Rats

To evaluate the effect of these compounds on thrombus formation in vivo,the following experimental protocol is performed.

Rats (CD-1; male; approximately 350 grams; Charles River, Raleigh,N.C.), are anesthetized with sodium pentobarbital (70 mg/kg i.p.). Theabdomens are shaved and a 22 gauge intravenous catheter is inserted intoa lateral tail vein. A midline incision is made and the intestines arewrapped in saline-soaked gauze and positioned so the abdominal aorta isaccessible. The inferior vena cava and abdominal aorta are carefullyisolated and a section (approximately 1 cm) of the abdominal aorta(distal to the renal arteries proximal to the bifurcation) is dissected.All branches from the aorta in this section are ligated with 4-0 silksuture. A 2.5 mm diameter flow probe connected to a Transonic flow meteris placed on the artery and a baseline (pre-stenosis) flow is recorded.Two clips are placed around the artery decreasing the vessel diameter byapproximately 80%. A second baseline flow measurement is taken(post-stenosis) and the hyperemic response is tested. Animals are thentreated with either compound or saline intravenously via tail veincatheter. Thrombosis is induced five minutes after treatment by repeatedexternal compressions of the vessel with hemostatic forceps. Two minutespost-injury, the vessel compressions are repeated and a 10 minute periodof flow monitoring is started. Animals are monitored continuously for aminimum of the first ten minutes post-injury. After twenty minutes(post-injury), a flow measurement is repeated and the animals areeuthanized. The section of the aorta that includes the injured sectionis harvested and placed in 10% formalin for possible histologicevaluation.

Example 18 In vivo PK/PD Measurements Following Oral Administration

To evaluate the ability of these compounds to be absorbed orally and toinhibit platelet aggregation in vivo, the following experimentalprotocol is conducted.

Male Sprague-Dawley rats are anesthetized using an inhaled anesthetic. Acannula containing heparinized saline is introduced into the jugularvein for withdrawal of venous blood samples. Animals are allowed a48-hour recovery period prior to dose administration.

Either compound or vehicle is administered to each animal as an oralgavage. Blood samples are taken immediately prior to compoundadministration, and at up to 12 time points ranging from 15 min to 24hours following compound administration. HPLC-MS/MS is used to measurethe amount of compound and/or metabolite in the blood samples.

Example 19 Inhibition of Thrombus Formation in Anesthetized Dogs

To evaluate the effect of the compounds of this invention on dynamicthrombus formation in vivo, the following experimental protocol, similarto the method of J. L. Romson et al. (Thromb. Res. 17:841-853, 1980), isperformed.

Surgical Preparation and Instrumentation: Briefly, purpose-bred dogs areanesthetized, intubated and ventilated with room air. The heart isexposed by a left thoracotomy in the fifth intercostal space andsuspended in a pericardial cradle. A 2-3 cm segment of the leftcircumflex coronary artery (LCCA) is isolated by blunt dissection. Theartery is instrumented from proximal to distal with a flow probe, astimulation electrode, and a Goldblatt clamp. The flow probe monitorsthe mean and phasic LCCA blood flow velocities. The stimulationelectrode and its placement in the LCCA and the methodology to induce anocclusive coronary thrombus have been described previously (J. K.Mickelson et al., Circulation 81:617-627, 1990; R. J. Shebuski et al.,Circulation 82:169-177, 1990; J. F. Tschopp et al., Coron. Artery Dis.4:809-817, 1993).

Experimental Protocol: Dogs are randomized to one of four treatmentprotocols (n=6 per treatment group) in which the control group receivessaline intravenously and the three drug-treated groups are administeredcompound intravenously. Upon stabilization from the surgicalinterventions, dogs receive either saline or compound. Afterapproximately 30 minutes, an anodal current is applied to the LCCA for180 min. The number and frequency of cyclic flow variations (CFV) thatprecede formation of an occlusive thrombus are recorded. These cyclicphenomena are caused by platelet thrombi that form in the narrowed lumenas a result of platelet aggregation (J. D. Folts et al., Circulation54:365-370, 1976; Bush et al., Circulation 69:1161-1170, 1984). Zeroflow in the LCCA for a minimum of 30 minutes indicates a lack ofantithrombotic efficacy (L. G. Frederick et al., Circulation 93:129-134,1996).

Example 20 Inhibition of ADP-induced Aggregation by Compounds of theInvention: IC₅₀ Values Obtained from Washed Platelets

The IC₅₀ values for the following compounds as inhibitors of ADP-inducedaggregation of washed platelets were determined using the proceduresprovided in Example 14:

-   4-{2,2-Dimethyl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-isophthalic    acid: IC₅₀=6.59 micromolar.-   2-[6-[6-(3-Phenyl-ureido)-purin-9-yl]-2-(2-trifluoromethyl-phenyl)-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy]-nicotinic    acid IC₅₀=8.07 micromolar.-   2-{2-Naphthalen-2-yl-6-[6-(3-phenyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinic    acid IC50=18 micromolar.-   2-{6-[6-(3-Hexyl-ureido)-purin-9-yl]-2-phenyl-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinic    acid IC₅₀=2.15 micromolar.-   2-{6-[6-(3-Ethyl-ureido)-purin-9-yl]-2-phenylethynyl-tetrahydro-furo[3,4-d][1,3]dioxol-4-ylmethoxy}-nicotinic    acid IC₅₀=1.0 micromolar.-   1-{2-Benzyl-6-[6-(3-cyclopentyl-ureido)-purin-9-yl]-tetrahydro-furo[3,4-d][1,3]dioxole-4-carbonyl}-pyrrolidine-2-carboxylic    acid IC₅₀=0.75 micromolar.-   1-(9-{2-Benzyl-6-[(3-methylsulfonyl-ureido)-methyl]-tetrahydro-furo[3,4-d][1,3]dioxol-4-yl}-9H-purin-6-yl)-3-phenyl-urea    IC₅₀=15 micromolar.

Example 21 IC₅₀ values for representative compounds of the presentinvention

Platelet IC₅₀ data were determined using washed human platelets,according to the protocol of example 14. Agonist challenge (ADP)typically in the range of 1-5 μM. Data are presented in μM and are fromthe average of two experiments or more.

TABLE 1 PLATELET DATA IC50 (uM) Compound # Washed Platelets 1 6.6 14 8.119 18 24 2.15 26 1 36 0.75 65 0.728 69 0.441 70 0.397 107 0.057 1080.098 109 0.03 110 0.023 112 0.009 115 0.012 127 0.032 128 0.042 1300.057 158 8 183 0.376 189 0.243 191 0.512 210 0.163 233 0.503 239 0.086298 0.081

The data in Table 1 illustrates that a diverse set of compounds of thepresent invention show activity as antagonists of P2Y₁₂-mediatedplatelet aggregation. For example, compounds falling under Formula I (1,14, 19, 24, 26, and 36) show P2Y₁₂ antagonist activity, despite havingdiffering moieties at the 4′ and 2′/3′ (acetal) positions of the ribose,and/or 6 (urea) position of the base. Additionally, compounds fallingunder preferred Formula IV (65, 69, and 70) illustrate thatdiastereomerically-pure molecules containing a prolinamide residue atthe 4′ position, a phenyl or styryl acetal moiety at the 2′/3′ position,and an ethyl urea at the 6 position have improved potency relative tothe less preferred compounds previously listed as falling under FormulaI. Further, compounds falling under a more preferred Formula V (107,108, 109, 110, 112, 115, 127, 128, and 130) show dramatically betterpotency (IC₅₀ data below 0.1 and even below 0.01 for Compound 112)believed to be as a result of their containing a benzyl ether (with orwithout substituents falling under the definition of M) or2-nicotinylmethyl moiety at the 4′ position, in addition to thepreferred acetals and urea previously listed for preferred compoundsfalling under Formula IV.

Table 1 also shows that replacing the oxygen of the benzyl ether moietyand reducing the linking group between the ribose ring 4′ position andthe terminal phenyl ring falling under the definition of X and FormulaII to two total atoms instead of three (158) still can lead to activemolecules. Yet further, preferred compounds falling under Formula VII(183, 189, 191 and 210) show that P2Y₁₂ antagonist activity can beobtained by modification of the ribose 4′ position with simple, acyclic,non-aromatic moieties, in conjunction with the preferred styryl acetaland ethyl urea moieties previously described. Even further, preferredcompounds falling under Formula IX (233) can show P2Y₁₂ antagonistactivity when one of the linking groups at the 4′ position falling underthe definition of A and B in Formula I is a carbamate moiety. In anotherillustration, preferred compounds falling under Formula X (239) showpotent activity when part of the linking group between the 4′ positionof the ribose and the phenyl ring falling under the definition of moietyX in Formula I and Formula II is an amide. In yet another illustration,compounds falling under the definition of Formula XII (298) can beactive when the moiety defined by X in Formula I and Formula II is a5-membered heterocyclic ring. Taken together, Table 1 illustrates that awide variety of molecules falling under the definitions of FormulaeIV-XII can be useful as antagonists of P2Y₁₂ mediated plateletaggregation, and consequently potentially useful as therapeutics indiseases where inhibition of platelet aggregation would be beneficial.

The invention, and the manner and process of making and using it, arenow described in such full, clear, concise and exact terms as to enableany person skilled in the art to which it pertains, to make and use thesame. It is to be understood that the foregoing describes preferredembodiments of the present invention and that modifications can be madetherein without departing from the scope of the present invention as setforth in the claims. To particularly point out and distinctly claim thesubject matter regarded as invention, the following claims conclude thisspecification.

1. A method of treating diseases or conditions associated with increasedplatelet aggregation, comprising administering to a subject in needthereof a therapeutically effective amount of a compound of Formula III,or a pharmaceutically acceptable hydrate, solvate or salt thereof,wherein said amount is effective to inhibit platelet aggregation:

wherein R_(a)=R_(c)=H; R_(b) is selected from the group consisting of:hydrogen, C₁₋₈ alkyl, C₁₋₈ alkenyl, C₁₋₈ alkynyl, C₃₋₇ cycloalkyl, C₄₋₇cycloalkenyl, aralkyl, and aryl, where all rings or chains optionallybear one or more substituents; R_(d) and R_(d′) are independentlyselected from the group consisting of: H, C₁₋₈ alkyl, C₁₋₈ alkenyl, C₁₋₈alkynyl, and C₃₋₇ cycloalkyl; R_(e) is absent; R_(f) and R_(g) areindependantly selected from the group consisting of: —H, halogen, C₁₋₈alkyl, C₁₋₈ alkenyl, C₁₋₈ alkynyl, and C₃₋₇ cycloalkyl; R_(h) is H,alkyl, aryl, or aralkyl; G is O; J is carbon; A₁ is O; D is O; X₁ isselected from the group consisting of: N and C-M; and M is independentlyselected from the group consisting of: —H, halogen, —CF₃, C₁₋₈ alkyl,cyano, C₁₋₈ alkenyl, C₁₋₈ alkynyl, C₃₋₇ cycloalkyl, C₃₋₇ cycloalkenyl,—OH, saturated or unsaturated C₁₋₆ alkoxy, amino, —N-substituted amino,and N,N-disubstituted amino.
 2. The method according to claim 1, whereinsaid compound reversibly inhibits ADP-induced platelet aggregation. 3.The method according to claim 1, wherein said compound is administeredin combination with other antiplatelet and/or anticoagulant drugs. 4.The method according to claim 1, wherein said diseases associated withplatelet aggregation are disorders characterized by thrombosis, primaryarterial thrombotic complications of atherosclerotic disease, thrombosissecondary to vascular damage and inflammation, indications with adiffuse thrombotic/platelet consumption component, venous thrombosis,coronary arterial thrombosis, pathological effects of atherosclerosisand arteriosclerosis, chronic or acute states of hyper-aggregability,reocclusion of an artery or vein following fibrinolytic therapy.
 5. Themethod according to claim 4, wherein said thrombosis are unstableangina, or myocardial infarction; said primary arterial thromboticcomplications of atherosclerosis are thrombotic stroke, peripheralvascular disease, or myocardial infarction said thromboses secondary tovascular damage and inflammation are vasculitis, arteritis,glomerulonephritis or organ graft rejection; said indications with adiffuse thrombotic/platelet consumption component are disseminatedintravascular coagulation, thrombotic thrombocytopenic purpura,hemolytic uremic syndrome, heparin-induced thrombocytopenia,pre-eclampsia or eclampsia; said venous thrombosis are deep veinthrombosis, veno-occlusive disease, or hematological conditions; andsaid coronary arterial thrombosis is associated with unstable angina,coronary angioplasty or acute myocardial infarction.
 6. The methodaccording to claim 5, wherein said pathological effects ofatherosclerosis and arteriosclerosis are arteriosclerosis, acutemyocardial infarction, chronic stable angina, unstable angina, transientischemic attacks, strokes, peripheral vascular disease, arterialthrombosis, preeclampsia, embolism, restenosis or abrupt closurefollowing angioplasty, carotid endarterectomy, or anastomosis ofvascular grafts; said chronic or acute states of hyper-aggregability iscaused by DIC, septicemia, surgical or infectious shock, post-operativetrauma, post-partum trauma, thrombotic thrombocytopenic purpura, snakevenom or immune diseases.
 7. The method according to claim 1, whereinsaid conditions associated with increased platelet aggregation and/orplatelet activation are produced by the contact of blood with anartificial device.
 8. The method according to claim 7, wherein saidartificial device is a hemodialysis instrument, a paracorporealartificial lung and an extracorporeal membrane oxygenation device, aninternal implantable artificial heart, an apheresis instrument used toremove or isolate a specific component of the blood and returning theremaining blood components to a donor.
 9. The method according to claim1, wherein said administering is systemic administration of saidcompound to a subject.
 10. The method according to claim 9, wherein saidsystemic administration is oral administration.
 11. A method for invitro inhibiting the aggregation of platelets in blood or blood product,comprising the step of administering to the blood or blood product thecompound according to claim 1, or a salt, solvate, or hydrate thereof.12. A method of treating patients to inhibit platelet aggregation in areversible manner, comprising the steps of: (a) providing a patient inneed of rapid reversal of platelet aggregation inhibition; (b)administering a therapeutically effective amount of a compound ofFormula I according to claim 1 to the patient; (c) submitting thepatient to a procedure selected from the group consisting of:percutaneous coronary interventions, stent placement, balloonangioplasty, coronary atherectomy, coronary endarterectomy, carotidendarterectomy, thrombolytic theraphy, coronary or other vascular graftsurgery, and dialysis, (d) discontinuing the administering of saidcompound to the patient; and (e) allowing the amount of said compound inthe patient's blood to reduce to below an therapeutically effectiveamount.
 13. The method according to claim 1, wherein said conditions areassociated with procedures resulting in platelet aggregation, saidconditions are selected from the group consisting of thromboticcomplications of interventions to treat atherosclerotic disease,thrombotic complications resulting from surgical procedures, thromboticcomplications resulting from mechanically-induced platelet activation,shunt occlusion, thrombosis secondary to vascular damage andinflammation, platelet adhesion associated with extracorporealcirculation, platelet activation associated with extracorporealcirculation, thrombotic complications associated with thrombolytictherapy, thrombotic complications associated with coronary and otherangioplasty, and thrombotic complications associated with coronaryartery bypass procedures.
 14. The method according to claim 13, whereinsaid thrombotic complications of interventions to treat atheroscleroticdisease are associated with procedures of angioplasty, endartectomy, orstent placement; said surgical procedures are coronary revascularizationprocedures, vascular graft surgery, tissue salvage following surgical oraccidental trauma, or reconstructive surgery; said mechanical-inducedplatelet activation is caused by cardiopulmonary bypass resulting inmicrothromboembolism, platelet refractoriness, and thrombocytopenia;said thrombotic complications resulting from shunt occlusion areassociated with procedures of renal dialysis or plasmapheresis.
 15. Themethod according to claim 1, wherein said diseases are stroke or angina.16. The method according to claim 4, wherein said diseases associatedwith platelet aggregation are disorders characterized by thrombosis,primary arterial thrombotic complications of atherosclerotic disease,thrombosis secondary to vascular damage and inflammation, indicationswith a diffuse thrombotic/platelet consumption component, venousthrombosis, coronary arterial thrombosis.
 17. The method according toclaim 1, wherein R_(d)′ is C₁₋₄alkyl or C₃₋₆cycloaklyl.
 18. The methodaccording to claim 1, wherein R_(b) is aryl, optionally bearing one ormore substituents.
 19. The method according to claim 17, wherein saidaryl is phenyl, benzyl, or styryl.
 20. The method according to claim 1,wherein R_(b) is aralkyl, optionally bearing one or more substituents.21. The method according to claim 1, wherein R_(g)=R_(f)=R_(d)=H; R_(d′)is C₁₋₄ alkyl, C₃₋₆ cycloalkyl; M is independently selected from thegroup consisting of: —H, halogen, —CF₃, C₁₋₄ alkyl, C₁₋₄ alkoxy, cyano,or amino; R_(b) is phenyl, benzyl, or styryl, optionally substituted.22. The method according to claim 1, wherein said compound is


23. The method according to claim 22, wherein said compound is Compound58, 60, or 62.